Novel clock gene bmal2

ABSTRACT

The present invention provides novel clock proteins BMAL2 (Brain-Muscle-Arnt-Like protein2), which is crucial for the clock oscillation mechanism including photic-input pathway and output pathway, novel clock genes encoding the proteins, a screening method using the proteins to screen a promoter or a suppressor of the promoter transactivation, and the like.  
     Genes for cCLOCK, cPER2, cBMAL1 were isolated from the chicken pineal gland which is a material suitable for studying circadian clock, then cDNA encoding the novel clock protein cBMAL2 having homology with cBMAL1 was isolated and sequenced. Further, BMAL2 cDNAs in human, mouse and rat were isolated respectively from the human embryonic kidney cell line, the mouse mid brain and the rat early fibroblast, and sequences of these cDNAs were determined. BMAL2 forms a heterodimer with CLOCK or BMAL1, etc. and it also forms a homodimer.

TECHNICAL FIELD

[0001] The present invention relates to novel proteins BMAL (Brain-Muscle-Arnt-Like protein) 2 which are involved in circadian rhythm, their genes, and their use.

BACKGROUND ART

[0002] Life activity is connaturally accompanied with various cyclic changes ranging from the behavior at the individual level to the biochemical phenomena at the cellular level. These rhythmic life activities occurring at certain cycles are called biorhythm and a periodic length of these phenomena which are repeated in cycles is often close to a periodic fluctuation of the environment such as a year, a month or a day. Sleep-wake rhythm and hormonal-secretion rhythm for such as melatonin and the adrenal cortex hormone are among those representing circadian rhythms repeated by an approximately 24-hour cycle, a daily unit. The circadian rhythms as mentioned have been observed in almost all the biological species and tissues and are regulated by the biological clock (Annu. Rev. Physiol. 55, 16-54, 1993). The suprachiasmatic nucleus (SCN) in the vertebrate central nervous system, pineal gland, specific neuronal tissues such as retina, etc. are known as tissues conforming circadian rhythm (Science 203, 1245-1247, 1979, Science 203, 656-658, 1979, Proc. Natl. Acad. Sci. USA 76, 999-1003, 1979, Brain Res. 245, 198-200, 1982, Neuron 10, 573-577, 1993, Science 272, 419-421, 1996).

[0003] As in the case of the mammalian suprachiasmatic nucleus (SCN), non-mammalian vertebrate pineal glands produce melatonin in response to circadian rhythm and light stimuli and play a central role in the physiological circadian regulation (Science 203, 1245-1247, 1979, Science 203,656-658, 1979, Proc. Natl. Acad. Sci. USA 76, 999-1003, 1979, Proc. Natl. Acad. Sci. USA 77, 2319-2322, 1980, Proc. Natl. Acad. Sci. USA 80, 6119-6121, 1983, J. Neurosci. 9, 1943-1950, 1989). The oscillation mechanism of the above-mentioned circadian rhythm is said to be characterized by the system wherein oscillation occurs at the gene level, is then amplified at the cellular level and finally reaches the individual level (Cell 96, 271-290, 1999). Oscillation at the gene level is brought by a group of genes called clock genes. Recent studies on the rodent clock genes have revealed that the circadian oscillator genes in mammals are positive and negative elements which form the transcription/translation-based negative feedback loop (Cell 96, 271-290, 1999, Annu. Rev. Neurosci. 23, 713-742, 2000). In mice, the negative elements include three period gene homologs; Perl (Cell 90, 1003-1011,1997, Nature 389,512-516,1997), Per2 (Cell 91, 1055-1064, 1997, Neuron 19, 1261-1269, 1997, Genes Cells 3, 167-176, 1998) and Per3 (EMBO J. 17, 4753-4759, 1998, Neuron, 20, 1103-1110, 1998) and two cryptochrome homologs; Cryl and Cry2 (Cell 98, 193-205, 1999, Nature 398, 627-630, 1999).

[0004] As for positive elements, BMAL1, CLOCK and the like which are basic helix-loop-helix (bHLH)-PAS (Per-Arnt-Sim) transcription elements are known. A CLOCK-BMAL1 complex is known to activate transcription through an E-box sequence (E-box: CACGTG) which is found not only in the negative element Perl (Science 280, 1564-1569, 1998) but also in clock-controleed genes such as vasopressin (Cell 96, 57-68, 1999) and in the albumin D-site binding protein gene (Genes Dev.14, 679-689, 2000). When a protein level of a negative element mentioned above is increased, its own transactivation for a promoter induced by a positive element is suppressed, the mRNA and protein levels of the negative element are down-regulated, and the molecular cycle is recommenced concomitant with the transactivation of the negative element gene. Therefore, the protein and mRNA levels of a negative element display a marked circadian oscillation. In addition to fluctuations in these clock genes, Perl and Per2 expressions are induced by light (Cell 91, 1055-1064, 1997, Neuron 19, 1261-1269, 1997, Cell 91, 1043-1053, 1997) and at least photo synchronization of an oscillatorisinducedbyPerl (J. Neurosci. 19,1115-1121,1999). Further, it has been revealed that mRNA levels of a positive element Bmall also exhibit circadian oscillation in antiphase to those of negative elements (Biochem. Biophys. Res. Commun. 250, 83-87, 1998, Biochem. Biophys. Res. Commun. 253, 199-203, 1998). Since its transcriptional rhythm is close to that of the Drosophila dClock (Science 286, 766-768, 1999), Bmall is thought to be involved in feedback loop of the negative elements (Science 286, 2460-2461, 1999, Science 288, 1013-1019, 2000).

[0005] On the other hand, the chicken (chick) pineal gland has been known that it retains the circadian oscillator as well as photic-input pathway and melatonin-output pathway in the pineal cell and that these properties can readily be retained under culturedconditions (Science 203, 1245-1247, 1979, Science 203, 656-658, 1979, Proc. Natl. Acad. Sci. USA 77, 2319-2322, 1980, Brain Res.438, 199-215, 1988, Recent Prog. Horm. Res. 45, 279-352, 1989, Nature 372, 94-97, i994, Proc. Natl. Acad. Sci. USA 94, 304-309, 1997, Brain Res. 774, 242-245, 1997). On the basis of these observations, the chick pineal cell is thought to be a prominent model for the study of the vertebrate circadian clock systems at the cellular level (Recent Prog. Horm. Res. 45, 279-352, 1989).

[0006] It is known that the biological clock is an auto- oscillatory system which oscillates autonomically without any exogenous stimulation and which, at the same time, has a property of being reset by the exogenous light-stimulation. It is also known that the vertebrate biological clock (circadian clock) which autonomically oscillates in a period close to a day is driven by the auto-feedback-loop consisting of a negative element and a positive element. Many things, however, still remain unknown with regard to the molecular clock system and the like including photic-input and output pathways. The object of the present invention is to provide novel proteins BMAL2 (Brain-Muscle-Arnt-Like protein 2) crucial in the clock oscillation mechanism including photic-input and output pathways, genes encoding the proteins, a method for screening a promoter or a suppressor of the promoter transactivation using the proteins, and the like.

DISCLOSURE OF THE INVENTION

[0007] The present inventors have made a keen study to solve the object mentioned above, and isolated cCLOCK, cPER2 and cBMAL1 genes from the chicken pineal gland which is a material suitable for the study of circadian clock, and further isolated cDNA encoding the novel clock protein cBMAL2 which was homologous with cBMALl and sequenced it. The inventors have also isolated the human, mouse and rat BMAL2 cDNAs respectively from the human embryonic kidney cell line, the mouse mid brain and the rat early fibroblast and sequenced them. In the pull-down assay, these novel clock proteins BMAL2 were found to form heterodimers with CLOCK, BMAL1 or the like, and to form homodimers among themselves (BMAL2). Besides, in the luciferase assay, BMAL2 were observed not only to form heterodimers with CLOCK and activate transcription via E-box but also to form homodimers and bind to E-box to cooperatively suppress transcription. Here the present invention is accomplished.

[0008] The present invention relates to: DNA encoding a protein (a) or (b) below,

[0009] (a) a protein comprising an amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8,

[0010] (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8 and which has the BMAL2 activity (claim 1); DNA containing a base sequence shown by Seq. ID No. 1, 3, 5 or 7 or its complementary sequence and part or whole of these sequences (claim 2); DNA which hybridizes with DNA of claim 2 under a stringent condition and which encodes a protein having the BMAL2 activity (claim 3); DNA encoding a protein (a) or (b) below,

[0011] (a) a protein comprising an amino acid sequence shown by Seq. ID No. 10,

[0012] (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 10 and which has the BMAL2 activity (claim 4); DNA containing a base sequence shown by Seq. ID No. 9 or its complementary sequence and part or whole of these sequences (claim 5); DNA which hybridizes with DNA of claim 5 under a stringent condition and which encodes a protein having the BMAL2 activity (claim 6); DNA encoding a protein (a) or (b) below,

[0013] (a) a protein comprising an amino acid sequence shown by Seq. ID No. 12 or 14,

[0014] (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 12 or 14 and which has the BMAL2 activity (claim 7); DNA containing a base sequence shown by Seq. ID No. 11 or 13 or its complementary sequence and part or whole of these sequences (claim 8); DNA which hybridizes with DNA of claim 8 under a stringent condition and which encodes a protein having the BMAL2 activity (claim 9); DNA encoding a protein (a) or (b) below,

[0015] (a) a protein comprising an amino acid sequence shown by Seq. ID No. 16, 18 or 20,

[0016] (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 16, 18 or 20 and which has the BMAL2 activity (claim 10); DNA containing a base sequence shown by Seq. ID No. 15, 17 or 19 or its complementary sequence and part or whole of these sequences (claim 11); and DNA which hybridizes with DNA of claim 11 under a stringent condition and which encodes a protein having the BMAL2 activity (claim 12).

[0017] The present invention further relates to: a protein comprising an amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8 (claim 13); a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8 and which has the BMAL2 activity (claim 14); a protein comprising an amino acid sequence shown by Seq. ID No. 10 (claim 15); a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 10 and which has the BMAL2 activity (claim 16); a protein comprising an amino acid sequence shown by Seq. ID No. 12 or 14 (claim 17); a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 12 or 14 and which has the BMAL2 activity (claim 18); a protein comprising an amino acid sequence shown by Seq. ID No. 16, 18 or 20 (claim 19); a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 16, 18 or 20 and which has the BMAL2 activity (claim 20); and a peptide which comprises part of the protein of any of claims 13-20 and which has the BMAL2 activity (claim 21).

[0018] The present invention still further relates to: a fusion protein or a fusion peptide wherein the protein of any of claims 13-20 or the peptide of claim 21 is bound with a marker protein and/or a peptide tag (claim 22); an antibody which specifically binds to the protein of any of claims 13-20 or to the peptide of claim 21 (claim 23); the antibody according to claim 23, wherein the antibody is a monoclonal antibody (claim 24); a recombinant protein or peptide to which the antibody of claim 23 or 24 specifically binds and which has the BMAL2 activity (claim 25); a host cell comprising an expression system capable of expressing the protein of any of claims 13-20 or the peptide of claim 21 (claim 26); the host cell according to claim 26, wherein the host cell is further capable of expressing CLOCK and/or BMAL1 (claim 27); the host cell according to claim 26 or 27, wherein the expression system at least comprises a promoter having an E-box sequence (CACGTG) (claim 28); the host cell according to claim 28, wherein the promoter having an E-box sequence (CACGTG) is a promoter of Per gene, Tim gene, Cry gene, vasopressin gene or the albumin D-site binding protein gene (claim 29); a non-human animal which, on its chromosome, is deficient in the gene function to encode the protein of any of claims 13-20 or the peptide of claim 21 or which over-expresses the protein of any of claims 13-20 or the peptide of claim 21 (claim 30); and the non-human animal according to claim 30, wherein the non-human animal is a mouse or a rat (claim 31).

[0019] The present invention also relates to: a method for screening a promoter or a suppressor for the expression of the protein of any of claims 13-20/the peptide of claim 21 or a promoter or a suppressor of the Bmal2 activity, wherein a cell expressing the protein or peptide and a test substance are used (claim 32); the method for screening a promoter or a suppressor for the expression of the protein/peptide or a promoter or a suppressor of the Bmal2 activity according to claim 32, wherein the cell expressing the protein of any of claims 13-20 or the peptide of claim 21 is the host cell of any of claims 26-29 (claim 33); a method for screening a promoter or a suppressor for the expression of the protein of any of claims 13-20/the peptide of claim 21 or a promoter or a suppressor of the Bmal2 activity, wherein the non-human animal of claim 30 or 31 and a test substance are used (claim 34); an expression promoter of the protein of any of claims 13-20 or the peptide of claim 21, wherein the expression promoter is obtained by the screening method according to any of claims 32-34 (claim 35); an expression suppressor for the protein of any of claims 13-20 or the peptide of claim 21, wherein the expression promoter is obtained by the screening method according to any of claims 32-34 (claim 36); a promoter of the Bmal2 activity obtained by the screening method according to any of claims 32-34 (claim 37); and a suppressor for the Bmal2 activity obtained by the screening method according to any of claims 32-34 (claim 38).

[0020] The present invention further relates to: a method for screening a promoter or a suppressor for the promoter transactivation, wherein a cell which expresses the protein of any of claims 13-20 or the peptide of claim 21 and which contains a promoter having an E-box sequence (CACGTG) and a test substance are used (claim 39); the method for screening a promoter or a suppressor for the promoter transactivation according to claim 39, wherein the cell which expresses the protein of any of claims 13-20 or the peptide of claim 21 and which contains a promoter having an E-box sequence (CACGTG) is thehost cellof claim 28 or 29 (claim 40); amethodfor screening a promoter or a suppressor for the transactivation for a promoter having an E-box sequence (CACGTG) in the non-human animal of claim 30 or 31, wherein the non-human animal and a test substance are used (claim 41); a promoter of the promoter transactivation obtained by the screening method according to any of claims 39-41 (claim 42); a suppressor for the promoter transactivation obtained by the screening method according to any of claims 39-41 (claim 43); and a method for diagnosing diseases associated with the expression or the activity of BMAL2, wherein the DNA sequence encoding BMAL2 in a sample is compared with the DNA sequence encuding the protein of claim 13 or 14 (claim 44).

BRIEF DESCRIPTION OF DRAWINGS

[0021]FIG. 1 shows the amino acid sequence of cPER2.

[0022]FIG. 2 shows the results of the amino acid homologies in domains among cPER2 and three mouse PER proteins (mPERl-3).

[0023]FIG. 3 shows the comparison among the amino acid sequences of various BMALs.

[0024]FIG. 4 shows the results of the amino acid homologies in domains among various BMAL proteins.

[0025]FIG. 5 shows the phylogenetic tree of ARNT-BMAL proteins and their amino acid homologies with cBMAL2 or hBMAL2.

[0026]FIG. 6 shows the genomic structure of hBMAL2 gene of the present invention.

[0027]FIG. 7 shows the basic structure of mouse BMAL2 and rat BMAL2 of the present invention.

[0028]FIG. 8 shows the phylogenetic tree of the BMAL-ARNT family proteins.

[0029]FIG. 9 shows the results of the northern blotting for analyzing the expressions of cBmal2 and cBamll genes of the present invention.

[0030]FIG. 10 shows the results of time-course changes in mRNA levels of cBmal1, cBmal2, cPer2 and cClock in the chicken pineal glands of the individuals.

[0031]FIG. 11 shows the time-course changes in mRNA levels of cBmal1, cBmal2, cPer2 and cCLOCK in the cultured chicken pineal cells under LD or DD condition.

[0032]FIG. 12 shows the results of the daily fluctuations under LD condition in mRNA expressions of mPer2, mClock, mBmal1 and mBmal2 in the mouse suprachiasinatic nucleus.

[0033]FIG. 13 shows the results of light-dependent changes in mRNA expressions of cPer2, cBmal1 and cBmal2 in the chicken pineal glands.

[0034]FIG. 14 shows the results of the in vitro physical interactions among cBMAL2 of the present invention, cBMALl and cCLOCK proteins.

[0035]FIG. 15 shows the results of the binding between a E-box sequence and cBMALl-cCLOCK or cBMAL2-cCLOCK detected by an electrophoretic mobility shift assay (EMSA).

[0036]FIG. 16 shows the results of transcriptional regulation in the 293EBNA cells induced by cBMAL1, cBMAL2 and cCLOCK.

[0037]FIG. 17 shows the cPER2 effect on transactivation mediated by E-box sequences.

[0038]FIG. 18 shows the effect of overexpression of cBMALl or cBMAL2 on the melatonin-rhythms of the chicken pineal cells.

BEST MODE OF CARRYUNG OUT THE INVENTION

[0039] Proteins of the present invention are exemplified by novel proteins with BMAL2 activity including: human BMAL2 shown by Seq. ID No. 2, 4, 6 or 8; chicken BMAL2 shown by Seq. ID No. 10; mouse BMAL2 shown by Seq. ID No. 12 or 14; rat BMAL2 shown by Seq. ID No. 16, 18 or 20; a protein comprising an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6, 8, 10, 12, 14, 16, 18 or 20, and having BMAL2 activity; and the like. Here the BMAL2 activity is taken to mean an activity to form a heterodimer with a transcription-promoting element to promote transcription via E-box in the promoter of a clock oscillator gene, and to form a homodimer to bind to E-box to competitively suppress transcription. Any peptide comprising part of the above-mentioned proteins and having BMAL2 activity may serve as a peptide as an object of the present invention, however, a peptide having a basic helix-loop-helix (bHLH) structure or a PAS (Per-Arnt-Sim) domain is preferable. Proteins and peptides as objects of the present invention and the recombinant proteins and peptides to which the antibodies, specifically binding to these proteins and peptides, bind specifically may collectively be referred to as “the present proteins/peptides” hereinafter. The present proteins/peptides can be prepared in accordance with known methods base on their DNA sequence information or the like and there should be no limitation as to the origin of the proteins/peptides.

[0040] Any DNA may be an object of the present invention as long as the DNA encodes the present proteins/peptides mentioned above and the specific examples include DNA encoding human BMAL2 shown by Seq. ID No. 2, 4, 6 or 8, DNA encoding chicken BMAL2 shown by Seq. ID No. 10, DNA encoding mouse BMAL2 shown by Seq. ID No. 12 or 14, DNA encoding rat BMAL2 shown by Seq. ID No. 16, 18 or 20; DNA encoding a protein comprising an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6, 8, 10, 12, 14, 16, 18 or 20 and having BMAL2 activity; and DNA containing the base sequence shown by Seq. ID No. 1, 3, 5, 7, 9, 11, 13, 15, 17 or 19 or its complementary sequence and part or whole of these sequences. These can be prepared by known methods from, for instance, a gene library or cDNA library and the like of human, chicken, mouse, rat, etc., based on their DNA sequence information or the like.

[0041] DNA encoding a protein having BMAL2 activity of the interest which has the same effect as human BMAL2, chicken BMAL2, mouse BMAL2, rat BMAL2, etc. can be obtained by hybridization with various DNA libraries under a stringent condition by using as a probe the base sequence shown by Seq. ID No. 1, 3, 5, 7, 9, 11, 13, 15, 17 or 19 or its complementary sequence and part or whole of these sequences, and by subsequent isolation of DNA which hybridized with the probe. DNAs thus obtained are also within the scope of the present invention. One example of a hybridization condition for obtaining DNA of the present invention is hybridization at 42° C. and washing at 42° C. in a buffer solution containing 1×SSC, 0.1% SDS, and more preferable example is hybridization at 65° C. and washing at 65° C. in a buffer solution containing 0.1×SSC, 0.1% SDS. There are number of factors other than the temperature condition mentioned above that affect the hybridization stringency and those skilled in the art can actualize the same stringency as that for the hybridization referred to in the above by appropriately combining various factors.

[0042] Any fusion protein and fusion peptide may be used as a fusion protein and a fusion peptide for the present invention as long as the present proteins/peptides are bound with marker proteins and/or peptide tags. As for a marker protein, there is no limitation as long as it is a conventionally known marker protein and the specific examples include alkaline phosphatase, the Fc region of an antibody, HRP, GFP, etc. Conventionally known peptide tags including Myc tag, V5 tag, HA tag, His tag, FLAG tag, S tag, etc. are the specific examples of a peptide tag for the use in the present invention. Such fusion protein can be generated according to ordinary protocols and is useful for the following: purification of the various BHAL2 or the like by using affinity of Ni-NTA and His tag; detection of a protein which interacts with various BMAL2; quantification of an antibody against various BMAL2 or the like; and use as a laboratory reagent in this field of art.

[0043] Antibodies that specifically bind to the aforementioned proteins and peptides of the present invention can be particularly exemplified by immune-specific antibodies including monoclonal antibodies, polyclonal antibodies, chimeric antibodies, single-stranded antibodies, humanized antibodies, etc. These antibodies can be generated according to ordinary protocols by using the above-mentioned various BMAL2 proteins or the like, or part of these proteins as an antigen. However, monoclonal antibodies are more preferable than the other sorts of antibodies mentioned because of their specificity. Antibodies such as the monoclonal antibodies are useful not only for diagnosis and treatment, such as missile therapy, for the circadian rhythm sleep disorders or the like including delayed sleep phase syndrome, non-24-hour sleep-wake syndrome, advanced sleep phase syndrome, time zone change syndrome, shift work sleep disorder, etc, but for elucidating the molecular mechanism of the circadian oscillation system.

[0044] Antibodies of the present invention are created by administering to an animal (preferably non-human) the present proteins/peptides, their fragments containing epitopes, or the cells expressing the proteins/peptides on the membrane surface, according to the conventional protocols. The monoclonal antibodies can be prepared, for instance, by any optional method that provides antibodies produced by cultured materials of continuous cell line such as a hybridoma method (Nature 256, 495-497, 1975), atriomamethod, ahumanB-cellhybridomamethod (Immunology Today 4, 72, 1983), and an EBV-hybridoma method (MONOCLONAL ANTIBODIES AND CANCER THERAPY, pp. 77-96, Alan R. Liss, Inc., 1985).

[0045] The preparation method for a single chain antibody (U.S. Pat. No. 4,946,778) can be adopted to prepare single-stranded antibodies to, the present proteins/peptides of the present invention mentioned above. Transgenic mice, other mammals, etc. can be used for expressing humanized antibodies. Clones expressing the present proteins/peptides can be isolated/identified using the antibodies mentioned above, and their polypeptides can be purified by affinity chromatography. Antibodies to the present proteins/peptides or to peptides containing their antigenic epitopes can possibly be used for diagnosis and therapy for circadian rhythm sleep disorders or the like including delayed sleep phase syndrome, non-24-hour sleep-wake syndrome, advanced sleep phase syndrome, time zone change syndrome, shift work sleep disorder, etc, and are useful for elucidating the molecular mechanism of the circadian oscillation system. Furthermore, recombinant proteins or peptides to which these antibodies specifically bind are also covered by the present proteins/peptides of the present invention as described earlier.

[0046] The functions of the present proteins/peptides can be analyzed by using, for example, antibodies such as the aforementioned monoclonal antibodies labeled with fluorescent materials including FITC (Fluorescein isothiocyanate), tetramethylrhodamine isothiocyanate, etc., radioisotopes including ¹²⁵I, ³²p, ¹⁴C, ³⁵S, ³H, etc., or enzymes including alkaline phosphatase, peroxidase, β-galactosidase, phycoerythrin, etc., or fused with fluorescent proteins such as Green Fluorescent Protein (GFP), BFP, CFP, YFP, RFP, etc. to serve as fusion proteins. As for immunological detection methods using the antibodies of the present invention, RIA method, ELISA method, fluorescent-antibody method, plaque method, spot method, haemagglutination, Ouchterlony method, etc. are exemplified.

[0047] There is no particular limitation as to a host cell of the present invention as long as the host cell comprises an expression system capable of expressing the present proteins/peptides. However, a preferable host cell is such in which the genes encoding CLOCK and/or BMAL1 are incorporated so that the two proteins can be simultaneously expressed in the host cell. Even more preferably, the host cell is incorporated with a DNA fragment which at least contains a promoter having E-box sequence (CACGTG), e.g. promoters of Per gene, Tim gene, Cry gene, vasopressin gene, the albumin D-site binding protein gene, etc., or a promoter introduced with E-box sequence (CACGTG) or the like. Although there is no particular limitation as to the above-mentioned DNA fragment so far as the fragment contains a promoter having E-box sequence (CACGTG), it is preferable for readily detecting and measuring the promoter activity that the DNA fragment is linked with a reporter gene including chloramphenicol acetyltransferase (CAT) gene, luciferasegene, etc., ageneencodingafluorescent protein including a short-lived green fluorescent protein (dlEGFP), etc. or with a fusion of GFP gene and a clock oscillator gene, and the like, to the down-stream of the promoter. Further, as to a promoter introduced with E-box sequence (CACGTG), any promoter may be adopted as long as its promoter activity can be regulated by a promoting element including the present proteins/peptides, CLOCK, BMAL1, etc. or by a suppressing element including PER, TIM, CRY, etc. These promoters are exemplified by RSV promoter, trp promoter, lac promoter, recA promoter, APL promoter, 1 pp promoter, SPO1 promoter, SPO2 promoter, penP promoter, PHO5 promoter, PGK promoter, GAP promoter, ADH promoter, SRα promoter, SV40 promoter, LTR promoter, CMV promoter, HSV-TK promoter, etc., but the promoters will not be limited to these exemplifications alone.

[0048] The present proteins/peptides and genes such as CLOCK and BMAL1 can be introduced into host cells by methods described in many standard laboratory manuals such as a manual of Davis et al. (BASIC METHODS IN MOLECULAR BIOLOGY, 1986), of Sambrook et al. (MOLECULAR CLONING: A LABORATORY MANUAL, 2nd Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989) and the like. The methods include calcium-phosphate transfection, DEAE-dextran-mediated transfection, transvection, microinjection, cationic lipid-mediated transfection, electroporation, transduction, scrape loading, ballistic introduction, infection, etc. The examples of host cells include bacterial prokaryotic cells such as E. coli, Streptomyces, Bacillus subtilis, Streptococcus, Staphylococcus, etc., eukaryotic cells such as yeast, aspergillus, etc., insect cells such as Drosophila S2, Spodoptera Sf9, etc., animal cells such as L cell, CHO cell, COScell, HeLacell, C127 cell, BALB/c3T3 cell (includingmutant strains deficient in dihydrofolate reductase, tymidine kinase, etc.), BHK21 cell, HEK293 cell, Bowes malignant melanoma cell, etc. and plant cells or the like.

[0049] There is no limitation to an expression system as long as the expression system is capable of expressing the present proteins/peptides described above in a host cell and the examples include chromosome-, episome- and virus-derived expression systems, for instance, vectors derived from bacterial plasmid, yeast plasmid, papovavirus such as SV40, vaccinia virus, adenovirus, fowlpox virus, pseudorabies virus and retrovirus, and vectors derived from bacteriophage, transposon and from the combination of these two, e.g. vectors derived from genetic factors of plasmid and bacteriophage such as cosmid and phagemid. Such expression system is not only for raising the expression and it may contain a regulatory sequence to regulate the expression.

[0050] Host cells comprising the above-mentioned expression systems and the present proteins/peptides obtained by culturing the cells can be used in a screening method of the present invention as described below. Further, the known methods can be adopted to collect and purify the present proteins/peptides from the cell culture, where the methods include ammonium sulfate- or ethanol-precipitation, acid extraction, anion- or cation-exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxyapatite chromatography and lectin chromatography, and the high performance liquid chromatography is preferably used. As a column especially used for affinity chromatography, for instance, columns to which antibodies to the present proteins/peptides are bound are used, and when common peptide tags are added to the present proteins/peptides mentioned above, columns to which substances having affinity with the peptide tags are bound are used, in order to obtain the present proteins/peptides. The purification methods for the present proteins/peptides mentioned above may also be employed for peptide synthesis.

[0051] In the present invention, a non-human animal whose gene function to encode. the present proteins/peptides mentioned above is deficient on its chromosome means a non-human animal part or whole of whose gene on its chromosome encoding the present proteins/peptides is inactivated by gene mutations such as destruction, deletion, substitutions, etc. and thus whose function to express the present proteins/peptides is lost. Further, a non-human animal which over-expresses the present proteins/peptides is specifically represented by a non-human animal which produces larger amount of the present proteins/peptides than a wild-type non-human animal does. Specific examples of non-human animals in the present invention include non-human animals such as rodents including mice, rats, etc., osteichthyes such as zebra fish, medaka fish, etc., arthropods such as Drosophila, silkworm, etc., the non-human animals should not be limited only to these examples.

[0052] Homozygous non-human animals that are born according to Mendel's Law include the deficient type or the over-expressing type for the present proteins/peptides as well as their wild type littermates. By using the deficient type animals or the over-expressing type animals of these homozygous non-human animals together with their wild-type littermates at the same time, accurate comparative experiments can be actualized out on the individual level. Therefore in performing screening of the present invention described below, it is, preferable to use wild type non-human animals, i.e. animals of the same species as, or even better the littermates of, non-human animals whose gene function to encode the present proteins/peptides is deficient or over-expressed on their chromosomes, in parallel with the deficient or over-expressed type animals. The method of generating a non-human animal whose gene function to encode the present proteins/peptides is deficient or over-expressed on its chromosome is now explained in the following with reference to a BMAL2 knockout mouse and a BMAL2 transgenic mouse.

[0053] A mouse, for instance, whose gene function to encode BMAL2 protein is deficient on its chromosome, i.e. a BMAL2 knockout mouse is generated by the following steps. A gene encoding mouse BMAL2 is screened by using a gene fragment obtained by a method such as PCR from a mouse gene library. A gene thus screened which encodes mouse BMAL2 is subcloned with a viral vector or the like and is identified by means of DNA sequencing. Then whole or part of a gene encoding BMAL2 among this clone is substituted with a pMC1 neo gene cassette or the like and then a gene such as a diphtheria toxin A fragment (DT-A) gene, a herpes simplex virus tymidine kinase (HSV-tk) gene, etc. is introduced onto either or both of 5′- or 3′-end, and thus a targeting vector is constructed.

[0054] The targeting vectors thus constructed are linearlized and introduced into ES cells by electroporation or the like to cause homologous recombination. Among the homologous recombinants, ES cells in which homologous recombination have occurred are selected by the use of antibiotics such as G418, ganciclovir (GANC), etc. It is preferable to confirm whether the ES cells selected are the recombinants of the interest by Southern blotting or the like. A clone of the ES cells thus confirmed is microinjected into a mouse blastocyst and which blastocyst is placed back to the recipient mouse to generate a chimeric mouse. A heterozygous mouse can be obtained by intercrossing the chimeric mouse and a wild type mouse. By further intercrossing the heterozygous mice, a BMAL2 knockout mouse of the present invention can be generated. Whether the ability of expressing BMAL2 is lost in the BMAL2 knockout mouse is examined by Northern blotting upon isolating RNA from the mouse obtained by the above-described method, and by Western blotting or the like in which the BMAL2 expression in the mouse can be directly examined.

[0055] A BMAL2 transgenic mouse is created by the following steps. A promoter such as chicken β-actin, mouse neurof ilament, SV40, etc. and poly (A) such as rabbit β-globin, SV40, etc. or introns are fused with cDNA encoding BMAL2 derived from chicken, mouse, human, rat, etc., to construct a transgene. This transgene is microinjected into the pronucleus of a mouse fertilized egg. After the obtained egg cell is cultured, it is transplanted to the oviduct of the recipient mouse which was bred thereafter. Neonatal mice having the aforementioned cDNA were selected from among all the mice born and thus the transgenic mice are created. Neonatal mice having the cDNA can be selected by extracting crude DNA from the mice tails or the like and then by performing a dot hybridization method using a gene encoding the introduced BMAL2 as a probe and by PCR method or the like using a specific primer.

[0056] Genes or DNAs encoding the present proteins/peptides, the present proteins/peptides, fusion proteins in which the present proteins/peptides and marker proteins and/or peptide tags are bound, antibodies to the present proteins/peptides, host cells comprising expression systems capable of expressing the present proteins/peptides, CLOCK, BMAL1, etc., non-human animalswhose gene function to encode the present proteins/peptides is deficient on their chromosome, non-human animals which over-express the present proteins/peptides and the like make it possible to elucidate the molecular mechanism of the circadian oscillation system. In addition to that, these can be used to screen a promoter or a suppressor for expression of the present proteins/peptides, a promoter or a suppressor for the Bmal2 activity, and a promoter or a suppressor for the promoter transactivation of the clock oscillator genes or the like. Some among the substances obtained by these screening methods may possibly be used for therapy of the circadian rhythm sleep disorders or the like including delayed sleep phase syndrome, non-24-hour sleep-wake syndrome, advanced sleep phase syndrome, time zone change syndrome, shift work sleep disorder, etc.

[0057] As for a screening method for a promoter or a suppressor for expression of the present proteins/peptides, or for a promoter or a suppressor for the Bmal2 activity of the present invention, methods are exemplified that use: cells expressing the present proteins/peptides and a test substance; and a non-human animal deficient in a gene function to encode the present proteins/peptides on its chromosome or a non-human animal overexpressing the present proteins/peptides and a test substance. A screening method using cells expressing the present proteins/peptides and a test substance, as mentioned above, can be exemplified by a method wherein a test substance is made to contact or introduced into, for instance, the cells expressing the present proteins/peptides, e.g. cells obtained from wild-type non-human animals, host cells of the present invention, cells obtained from transgenic non-human animals of the present invention, etc. and wherein the Bmal2 activity and changes in the expression levels of the present proteins/peptides are measured and assessed, but the methods should not be limited to these examples alone.

[0058] As for a screening method wherein a non-human animal whose gene function to encode the aforementioned present proteins/peptides is deficient on its chromosome or a non-human animal which over-expresses the present proteins/peptides is used along with a test substance, the examples specifically include: a method wherein a non-human animal whose gene function to encode the aforementioned present proteins/peptides is deficient on its chromosome or a non-human animal which over-expresses the present proteins/peptides, as mentioned above, is administered with a test substance and subsequently the Bmal2 activity and changes in the expression levels of the present proteins/peptides in the cells obtained from the non-human animal are measured and assessed; or a method wherein a non-human animal whose gene function to encode the aforementioned present proteins/peptides is deficient on its chromosome or a non-human animal which over-expresses the present proteins/peptides mentioned above is administered with a test substance and subsequently the Bmal2 activity and changes in the expression levels of the present proteins/peptides in the non-human animal are measured and assessed.

[0059] An example of a screening method of the present invention for a promoter or a suppressor of the promoter transactivation is a method wherein a test substance and a cell expressing either the present proteins/peptides or the present proteins/peptides along with CLOCK and/or BMAL1 and containing a promoter which has E-box sequence(CACGTG), morespecifically a method in which a test substance is made to contact or introduced into the aforementioned cell and the promoter activity mediated by E-box is then measured and assessed. Another example is a method wherein a test substance is applied to a non-human animal whose gene function to encode the present proteins/peptides is deficient on its chromosome or to a non-human animal which over-expresses the present proteins/peptides to measure and assess the change in the promoter activity mediated by E-box. In addition, it is preferable to have reporter genes or the like, such as chloramphenicol acetyltransferase (CAT) gene or luciferase gene, linked to the downstream of a promoter having E-box sequence (CACGTG), in order to readily analyze the promoter activity.

[0060] The present invention also relates to a diagnostic method for diseases associated with the activity or expression of BMAL2 protein wherein the method comprises comparing the DNA sequence encoding BMAL2 protein in a sample with the DNA sequence encoding BMAL2 protein of the present invention. Mutants of DNA encoding BMAL2 protein can be detected by finding individuals with gene mutations at the DNA level, and such detection is effective for diagnosing with diseases developed by underexpression, overexpression or mutated expression of BMAL2 protein. Specific examples of samples used for the detection include cells of a subject, for example, genomic DNA obtainable biopsy of blood, urine, saliva, tissue, etc., or RNA or CDNA. The samples, however, should not be limited to these exemplifications and the amplified products of PCR or the like may also be employed in using the samples. Deletions or insertion mutations of a base sequence can be detected through the changes in size of the amplified products when compared to that of the normal gene type. Point mutation can be identified by hybridizing the amplified DNA with a gene encoding a labeled BMAL2 protein. As described. in the above, the circadian rhythm sleep disorders or the like including delayed sleep phase syndrome, non-24-hour sleep-wake syndrome, advanced sleep phase syndrome, time zone change syndrome, shift work sleep disorder, etc. can be diagnosed or judged by detecting mutation in a gene which encodes BMAL2 protein.

[0061] The present invention is now further described specifically with reference to the examples, however, the scope of the invention should not be limited to these examples alone.

EXAMPLE 1 Cloning and Sequencing

[0062] 1-1 (Cloning and sequencing of cClock cDNA)

[0063] cClock cDNA was amplified with the chicken pineal cDNA library (AZAPII, 5×10⁵ pfu) as a template by PCR using LA-Taq polymerase (Takara) and a pair of primers [sense primer 1: 5′-ACTAGTCGACTTATGTTTTTTACCATAAGCACC-3′ (Seq. ID No. 21), antisense primer 1: 5′-GTCGACCTGCGCTACTGTGGCTGAGCTTTG-3′ (Seq. ID No. 22); Each of the primers has a SalI site on its 5′-terminal] which were designed on the basis of the sequences of cClock genes deposited in GenBank (GenBank accession nos. AF132531 and AF144425). The above-mentioned PCR method was performed four different times and the sequences of the five clones obtained were determined. One clone with no PCR error was selected (GenBank accession no. AF246959). The program for thermal cycles was as follows: degeneration for 1 min at 94° C. only for the first time, followed by 5 repetitive cycles each consisting of thermal degeneration for 30 sec at 94° C., annealing for 30 sec at 55° C. and extension for 3.5 min at 72° C.; followed by 15 repetitive cycles each consisting of thermal degeneration for 30 sec at 94° C., annealing fur 30 sec at 65° C. and extension for 3.5 min at 72° C.; and finally extension for 6.5 min at 72° C.

[0064] 1-2 (Cloning and sequencing of cPer2 cDNA)

[0065] A 273 bp fragment of cPer2 cDNA was obtained from a chicken pineal cDNA library by PCR using Taq-Gold polymerase (PE applied biosystems) and a pair of degenerate primers [per-F, 5′-CAGCAGAT(C/G)A(A/G)CTG(C/T)IT(C/G)IGACAG(C/T)(A/G)TC(A/C)TCAG-3′ (Seq. ID No. 23) and per-R, 5′-GCT(A/G)CACTG(A/G)CTG(A/G)TG(A/C)(C/G)IGAC(A/G)CCAC(A/G)CTC-3′ (Seq. ID No. 24)] which were designed based on the nucleotide sequences of dPer and mammalian Per genes. A longer cDNA fragment (P2-5; 886bp) was amplified from a chicken pineal cDNA library by the subsequent PCR using cPer2-Rl primer [5′-TTGCTGTACCAGGCACATTACAAC-3′ (Seq. ID No. 25)] synthesized from the base sequence of the above-obtained fragment, a degenerate primer [YK-F1; 5′-(A/G)TICA(C/T)TCIGGITA(C/T)CA(A/G)GCICCI(A/C)GIATICC-3′ (Seq. ID No. 26)] and LA-Taq polymerase. This fragment was used as a hybridization probe for the screening of the chicken pineal cDNA library (λZAPII, 5×10⁵ pfu) to isolate a clone Pa (3584 bp) encoding a larger part of cPER2 (Met¹-Arg¹⁰¹⁴). This clone and the cDNA clone obtained by 3′-RACE were ligated together to generate a full-length clone for cPER2 (Met¹-Thr ¹³⁴⁴; GenBank accession no. AF246956). The result is shown in FIG. 1 in which the DNA sequence and the amino acid sequence are shown as Seq. ID Nos. 27 and 28 respectively. The bars above the sequence in FIG. 1 indicate the PAS domains (PAS-A and PAS-B) and the cytoplasmic localization domain (CLD). FIG. 2 shows the amino acid homologies in domains between cPER2 obtained as above and three mouse PER proteins (mPER1-3). The programming for thermal cycles of the above was as follows: degeneration for 1 min at 94° C. only for the first time; followed by 35 repetitive cycles each consisting of thermal degeneration for 30 sec at 94° C., annealing for 60 sec at 52° C. and extension for 1 min at 72° C.; and finally extension for 9 min at 72° C.

[0066] 1-3 (Cloning and sequencing of cBmal cDNA)

[0067] cDNA clones encoding part of cBMAL1 or cBMAL2 were respectively obtained from the chicken pineal cDNA library by PCR using LA-Taq polymerase with degenerate primers [BMAL-F, 5′-GTGCT(A/C)(A/C)GGATGGC(A/T)GT(G/T)CAGC-3′ (Seq. IDNO. 29) and BMAL-R, 5′-GCG(C/T)CC(A/G)ATTGC(A/C/G)AC(A/G)AGGCAG-3′ (Seq. ID No. 30)] which were designed based on nucleotide sequences of Bmall of mouse, rat and human and dcycle of Drosophila. Each amplified fragment and a cDNA clone of the each amplified fragment obtained by 5′-RACE were used as probes for screening the chicken pineal cDNA library (AZAPII, 3.5×10⁵ pfu) and cDNA clones containing the coding regions for cBMAL1b′ (GenBank accession no. AF246957) and cBMAL2 (GenBank accession no. AF246958) were respectively isolated and sequenced (FIG. 3). The bars above the sequences in FIG. 3 indicate the basic helix-loop-helix region (bHLH) and PAS domains (PAS-A and PAS-B). PCR for the above was performed using a thermal cycler (Perkin-Elmer) as follows: thermal degeneration for 1 min at 94° C. only for the first time; followed by 35 repetitive cycles each consisting of thermal degeneration for 30 sec at 94° C., annealing for 1 min at 50° C. and the extension reaction for 1 min at 72° C.; and finally extension for 9 min at 72° C.

[0068] The initiation methionine of cBMAL1b′ was predicted by comparing the cBMALlb′ sequence mentioned above and the BMAL1 sequences of other animal species. The initiation methionine of the aforementioned cBMAL2 was predicted by the following three criteria; (i) A nonanucleotide sequence (CCGCCATGG), the 97-105 base sequence of cBmal2 shown as Seq. ID No. 9, fully matched the Kozak's translation initiation consensus sequence (Nucleic Acids Res. 12. 857-872, 1984). (ii) The above-mentioned Bmal2 cDNA clone (3.4 kb) and mRNA (3.0, 3.8 kb) were similar in size to each other. (iii) A promoter region predicted from its genomic analysis contained the upstream inframe stop codons.

[0069] Next, the amino acid homologies in domains among mBMALlb′ and three novel BMAL proteins (cBMAL1b′, cBMAL2 and hBMAL2a) were analyzed and a phylogenetic tree of ARNT and BMAL proteins was constructed according to Neighbor-joining method using PHYLIP, v.3.572 as described in the literature (Felsenstein, J., PHYLIP, Version 3.572, University of Washington, Seattle, 1996). These results are respectively shown in FIGS. 4 and 5. In FIG. 5, since amino acids in cBMAL2 in the amino-terminal region (Met¹-Arg¹⁰⁴) and carboxy-terminal region (Gly⁴⁵⁹-Leu⁶²²) differ in number among animal species, a part corresponding to this region was omitted from each protein before calculating the amino acid homologies among the proteins, and then the phylogenetic tree was constructed. These results demonstrate that cBMAL1b′ is 93% homologous to mBMAL1b′ to show they are close to one another (FIGS. 3 and 4), while cBMAL2 (ARNT4) is not particularly close to BMALL (70%; FIG. 5) nor to ARNT1 (41%; FIG. 5) nor to ARNT2 (40%; FIG. 5) and hence that cBMAL2 is a novel protein having bHLH-PAS (FIG. 4).

[0070] 1-4 (Cloning and sequencing of hBmal2 cDNA)

[0071] A partial sequence information of hBmal2 was obtained from two human EST clones (GenBank accession nos. AA577389 and AI218390) by in silico screening using cBmal2 as a probe (data as of October 1999). Several cDNA clones containing the 5′-untranslated region of hBmal2 gene were isolated from cDNA of 293EBNA cell (a human embryonic kidney cell line) by 5′-RACE. Then full-lengthclones were amplifiedbyPCRusinghB2F1 andhB2Rlprimers [hB2F1, 5′-GACCAAGTGGCTCCTGCGAT-3′ (Seq. ID No. 31) and hB2R1, 5′-GCTAGAGGGTCCACTGGATG (Seq. ID No. 32)]. To eliminate PCR errors, 17 full-length cDNA clones obtained were sequenced, and all the DNA sequences encoding hBMAL2a- d (GenBank accession nos. AF246960-AF246963), which were consistent with the human genomic sequences (GenBank accession nos. AC021737 and AC016008), were determined. The programming for the PCR thermal cycles mentioned above was as follows: degeneration for 1 min at 94° C. only for the first time; followed by 20 repetitive cycles each consisting of thermal degeneration for 30 sec at 94° C., annealing for 60 sec at 60° C. and extension for 2 min at 72° C.; and finally extension for 8 min at 72° C. These results are shown in FIGS. 3, 4 and 5. The arrowheads below the sequences in FIG. 3 indicate the insertion sites of introns in hBmal2 gene.

[0072] cDNA sequences encoding 4 variants of hBAML2 (hBMAL2a-d) and obtained from 293EBNA cells as described above, were compared with the genome sequences registered at GenBank (accession nos. AC021737 and AC016008). Then the cDNA sequences were divided into 17 exons as in the case of mBmall (Biochem. Biophys. Res. Commun. 260, 760-767, 1999) to examine the genomic organization of hBmal2. The results are shown in FIG. 6. Bars with GenBank accession numbers in FIG. 6 represent genomic and CDNA clone regions and shaded parts are the spliced regions in the isolated mutants. These results show that the cDNA clone of hBMAL2b is devoid of Exon 4 (corresponding to Val⁹⁶-Arg¹⁰⁹ in hBMAL2a) and that of hBMAL2c is devoid of both Exons 3 and 4 (corresponding to Gln⁷⁵-Arg¹⁰⁹ in hBMAL2a) and having Exon 1 to which DNA encoding the amino acid sequence of 11 amino acid residues (GEVAGGEATAP) added in-between Gly¹⁰ and Gly¹¹ in hBMAL2a is extended. hBMAL2d was revealed to be the shortest mutant which is devoid of both Exon 1 (as in hBMAL2a/b) and Exons 3/4 (as in hBMAL2c) in cDNA.

[0073] 1-5 (Cloning and sequencing of mBmal2 cDNA)

[0074] To identify the mouse Bmal2 ortholog (mBmal2) expressed in the suprachiasmatic nuclei (SCN), a 629 bp fragment CDNA was obtained by RT-PCR for total RNA extracted from the mouse mid-brain, by using LA-Taq polymerase (Takara) and two primers synthesized according to the hBmal2 sequence; [hBMAL2-F4: 5′-GTGCTGGTAGTATTGGAACAGATATTG-3′ (Seq. ID No. 33) and hBMAL2-R1: 5′-GCTAGAGGGTCCACTGGATG-3′ (Seq. IDNo. 34). Subsequently, several cDNA clones were isolated which contain 5′- or 3′- untranslated region of mBmal2 cDNA by the method of 5′- and 3′- rapid amplification of cDNA ends. Based on these sequence information, two primers [mBMAL2-F1: 5′-GGTCGACCACCATGGAGTTTTCCAAGGAAACG-3′ (Seq. ID No. 35), mBMAL2-R1: 5′-GCTAGAGTGCCCACTGGATGTCAC-3′ (Seq. ID No. 36)] were designed that were capable of amplifying full-length clones covering the total coding sequence of mBMAL2a or mBMAL2b (FIG. 7; GenBank accession nos. AY005163 and AY014836). Another RT-PCR was performed using these primers and LA-Taq polymerase to obtain mBMAL2a comprising 579 amino acid residues. This amino acid sequence contained bHLH, PAS-A and PAS-B domains and was homologous to hBMAL2 by 74%, cBMAL2 by 63% and zBMAL2 by 48%. On the contrary, mBMAL2b consists of amino acid residues that are about one third of those of mBMAL2a (199 amino acid residues) and is devoid of PAS-B domain (FIG. 7). Although this form of mutation is similar to that previously found in hBMALlc (a BMAL1 mutant devoid of the C-terminal half in the BMAL1 comprising a long chain; Biochem. Biophys. Res. Commun. 233, 258-264, 1997), its physiological meaning is yet unknown.

[0075] 1-6 (Cloning and sequencing of rBmal2 cDNA)

[0076] Next, cDNA clone of rat Bmal2 (rBmal2) covering almost a total coding region was isolated from the rat early fibroblast rat-1 cells by RT-PCR using two primers [mBMAL2-Fl and mBMAL2-R1]. Three clones isolated, rBMAL2a-c, were determined for the amino acid sequences (FIG. 7; respectively registered to GenBank under GenBank accession nos. AF327071, AY014837, AY014838). The amino acid sequence at the amino-terminal in rBmal2 (corresponding to the position of mBMAL2-Fl primer) was obtained from the in silico screening (GenBank accession no. AA944306). These results demonstrate that rBMAL2a comprising the longest sequence among the clones obtained is most similar to mBMAL2a in its structure. In FIG. 7, dots at the end of the rBMAL2 sequence indicate the position corresponding to mBMAL2-R1, a PCR primer. The asterisk indicates the position of the in-frame stop codon of mBMAL2b and the number at the end of each line (with “+” on the right shoulder) indicates the number of amino acid residues for rBMAL2a.

[0077] Next, the phylogenetic tree for the BMAL-ARNT family was constructed according to the amino acid homologies among various proteins (FIG. 8). Before constructing the phylogenetic tree, several amino acid sequences of BMAL-ARNT proteins that were obtained from GenBank were aligned with Gene Works (Ver.2.55, clustal V), then some regions with gaps were omitted. Since the length of amino acids in amino- and carboxyl-terminal regions (corresponding to the 1-59 amino acid sequence and the 413-579 amino acid sequence of mBMAL2a) differ among mutants, these regions were also omitted. Then the Neighbor-joining tree was constructed using a PHYLIP 3.572 software package (Felsenstein, J., PHYLIP, Version 3.572, University of Washington, Seattle, 1996) (FIG. 8), and the topology of the phylogenetic tree obtained as above was analyzed by PROTML 2.3 program which adopts a local rearrangement method for the maximum likelihood analysis and JTT-F model for the amino acid substitution (Adachi, J. and Hasegawa, M., MOLPHY: Programs for molecular phylogenetic based on maximum likelihood, Version 2.3, Institute of Statistical Mathematics, Tokyo, 1996). Further, in order to assess the reliability of that phylogenetic tree, a boot strap test was carried out and the boot strap probabilities of over 70% were respectively shown near the diversion points in FIG. 8. The diversion points shown by closed circles indicate the divergence of species and those shown by open circles indicate gene duplications in FIG. 8.

[0078] When the above result is taken into account together with the fact that there is only a single copy of dcyc gene, a Bmal1/2-like gene, in the Drosophila genome, Bmall and Bmal2 genes are likely to be generated from the gene duplication occurred in their ancestral vertebrates (Diversion point b in FIG. 8). Besides, branches at the divergence among the members in the BMAL2 cluster are much longer than those of BMAL1, meaning that the phylogenetic tree topology in the BMAL2 cluster reflects the phylogenetic development of vertebrates. It can therefore be concluded from these facts that these Bmal2 genes are in orthologous relationships with each other and have developedfromahighlyfrequent aminoacidsubstitution. This conclusion can also be supported by the fact that no m/r/c/z Bmal2 orthologs other than hBmal2 can be found in the human gene data base (the htgs database was searched on 9^(th) Dec., 2000). Diversion point a in FIG. 8 probably indicates divergence between ancestors of vertebrates and arthropods and diversion points c-f indicate divergence among vertebrate species. Besides, the above-mentioned phylogenetic tree had the same topology as phylogenetic trees obtained by Parsimony and Neighbor-joining methods.

[0079] Comparison of substitution rates in amino acids among the members of BMAL1/2 clusters revealed that the amino acid substation rate of BMAL2 is higher than that of BMAL1 by about 20-fold. This shows that the selective pressure in BMAL2 after gene duplication is lower than that in BMAL1. What is important is that there is no any specific region in which the total amino acid homology among BMAL2 proteins is decreased. Highly conserved structure of BMAL1 protein which has a higher selective pressure is thought to include some unrecognized function which has been lost in BMAL2. BMAL1 is thought to interact with several essential regulatory factors that have not yet been characterized, because both BMAL proteins interact with CLOCK which is a functional heterodimer partner with BMAL proteins (Science 280, 1564-1569, 1998, Proc. Natl. Acad. Sci. USA 97, 4339-4344, 2000, J. Neurosci. 20, RC83, 2000, J. Biol. Chem. 275, 36847-36851, 2000, Proc. Natl. Acad. Sci. USA 95, 5474-5479, 1998, Biochem. Biophys. Res. Commun. 248, 789-794, 1998), with a neuron PAS domain protein 2 (NPAS2 or MOP4) (J. Neurosci. 20, RC83, 2000, Proc. Natl. Acad. Sci. USA 95, 5474-5479, 1998), with a hypoxia-inducing factor la (HIFla) (J. Neurosci. 20, RC83, 2000, Proc. Natl. Acad. Sci. USA 95, 5474-5479, 1998, Biochem. Biophys. Res. Commun. 248, 789-794, 1998),or with HIF2α(HLF or EPAS1) and with the like. Therefore, analyzing the differences between the functions of BMAL1 and BMAL2 is thought to contribute to uncover their unique evolution processes.

EXAMPLE 2 Northern Blot Analysis

[0080] Total RNA (7.5 μg) of each tissue from one-week-old chicks (pineal gland, retina, cerebrum, heart, kidney and skeletal muscle) was analyzed by Northern blotting in a manner as described in J. Neurochem. 70, 908-913, 1998. These tissues were harvested at 0, 6, 12 and 18 hr in Zeitgeber time (ZT), frozen with liquid nitrogen and mixed before extracting RNA. Each of total RNA was separated by an agarose gel electrophoresis and blotted on a nitrocellulose membrane. The blotting membrane was hybridized with a cBmall probe or a cBmal2 probe and washed (10 min×3 times) in 0.1×SSC at 50° C., then analyzed using a FLA2000 bioimage analyzer (FUJI PHOTO FILM). The membrane was subsequently hybridized with a chicken histone H4cDNA probe and analyzed. The chicken histone H4cDNA probe used was prepared by amplification by PCR with a primer [sense primer 2; 5′-CATGTCTGGCAGAGGCAAG-3′ (Seq. ID No. 37) and antisenseprimer2; 5′-TTAGCCGCCGAAGCCGTAG-3′ (Seq. IDNo.38)], which was designed from the chicken pineal cDNA based on the sequence (accession no. M74533) deposited in GenBank, and by the subsequent cloning. The results are shown in FIG. 9. These results demonstrate that two cBmal2 genes (3.8 Kb and 3.0 Kb, indicated by arrows) and cBmall gene (3.3 Kb) are expressed in all the tissues examined at various intensities. It was confirmed as a result of normalization to histone H4 that heart and kidney exhibited low transcriptional levels of cBmall and that no apparent difference was observed in the transcriptional levels of cBmal2 among the tissues examined.

EXAMPLE 3 Expression of Chicken Clock Genes in the Pineal Gland

[0081] One-day-old chicks were entrained to LD cycle (12 hr with light/12 hr in the dark) for 3 weeks, then placed in DD (constant darkness) condition for 2 days, and the pineal glands were collected every 4 hr over the last 3 days. Total RNA from each pineal gland was analyzed by Northern blotting to detect expression of chicken Clock genes (cBmall, cBmal2, cPer2 and cClock) in the pineal gland. Total RNA (6 μg) obtained from each pineal gland mentioned above was separated by an agarose gel electrophoresis, blotted on a nitrocellulose membrane. Two such blotting membranes were prepared. A blot was first hybridized with a cBmal2 probe or a cPer2 probe and the blotting membrane was washed in 0.1×SSC at 50° C. (10 min×3 times), which was then analyzed using a FLA2000 bioimage analyzer (FUJI PHOTO FILM). Next, the blot was hybridized with the histone H4cDNA and analyzed in the same way. The aforementioned cDNA fragment P2-5wasusedas thecPer2 probe. Foranotherblotting membrane, the blot was first hybridized with a cBmall probe as in the above, then with a histone H4cDNA and finally with a cClock probe, and was analyzed with a FLA2000 bioimage analyzer. These results are shown in FIG. 10 (bottom lane). Signals for cBmall (open circles) and cBmal2 (closed circles) were quantified by MacBAS software (FUJI PHOTO FILM), normalized to those for the histone H4 cDNA, and the mean value was set as 1 in each case to analyze the time-course changes in transcriptional levels of the chicken Clock genes. The results are shown in FIG. 10 (top lane). A cross bar above the Northern blotting results in FIG. 10 indicates light and bright cycles. An open region indicates a light cycle, closed regions indicate (subjective) dark cycles and shaded regions indicate subjective light cycles. Three cPer2 transcripts (9.7 Kb, 7.5 Kb and 4.1 Kb) and a single cClock transcript (8.5 Kb) were confirmed by these results.

EXAMPLE 4 Expression of Chicken Clock Genes in the Pineal Cell Culture

[0082] The time course changes in the transcription amounts of chicken clock genes [cBmall (open circle), cBmal2 (closed circle), cPer2 (open triangle) and cClock (open square)] in the pineal cell culture were analyzed by a quantitative RT-PCR method and the results were compared to those in Example 3 above (FIG. 11). Pineal cells from one-day-old chicks were plated on 35 mm dishes (cells from 8 pineal glands per a dish) and cultured for 5 days under LD cycles in Medium 199 (Life Technologies) supplemented with 10% fetal bovine serum. On day 6, part of the cultured cells was shifted to culture under constant darkness (DD, right in FIG. 11). The rest of the cultured cells remained in the culture under the LD condition and subjected to a further culture on day 7 under constant darkness (LD, left in FIG. 11). Then each pineal cell was harvested every 4 hours. The pineal cells harvested were suspended in TRIzol reagent (Life Technologies) and stored at −80° C. until total RNA was isolated. 1 μg each of the total RNA was reverse-transcribed by the SuperScriptII (Life Technologies) reverse transcriptase and a portion of the reaction product was used for PCR analysis. First, an optimal number of PCR cycle was determined for each primer to give linear relationships between the amounts of the template cDNA and amplification products and PCR was carried out under such condition. The PCR products obtained were separated by a 7.5% polyacrylamide gel electrophoresis, stained with SYBR Green I (Molecular Probes), and the transcriptional level of each chicken clock gene was quantified with a FLA2000 bioimage analyzer (FUJI PHOTO FILM). Change in the transcriptional level of GAPDH, as a control, was measured in a similar manner as the above. Intensity of each signal was normalized to that of GAPDH, and the mean value for each gene on day 6 was set to 1. Then all the values (mRNA levels) were obtained from three different culture samples, which were shown by mean±SEM.

[0083] The primers and number of PCR cycles mentioned above were set up as follows. For cBmall, cBlF1600 primer; 5′-TCCAGACATTTCTTCAGCTGG-3′ (Seq. ID No. 39) and cBIREND-primer; 5′-GGATGTTGAAGCAAGGTGC-3′ (Seq. ID No. 40) were used and 23 cycles were practiced. For cBmal2, cB2F1270-primer; 5′ACGAGTACTGCCATCAAGATG-3′ (Seq. ID No. 41) and cB2REND-primer; 5′-GAGAGCCCATTGGATGTCAC-3′ (Seq. ID No. 42) were used and 23 cycles were practiced. For cClock, cqCF862-primer; 5′-TTCTTGGATCACAGGGCAC-3′ (Seq. ID No. 43) and cqCR1364-primer; 5′-GGAGTGCTAGTGTCCACTGTCA-3′ (Seq. ID No. 44) were used and 25 cycles were practiced. For cPer2, cP2RTF primer; 5′-GGAAGTCCTTGCAGTGCATAC-3′ (Seq. ID No. 45) and cP2RTR-primer; 5′-ACAGGAAGCGGATATGCAG-3′ (Seq. ID No. 46) were used and 24 cycles were practiced. For GAPDH (GenBank accession no. K01458), cGAF-primer; 5′-ACCACTGTCCATGCCATCAC-3′ (Seq. ID No. 47) and cGAR-primer; 5′-TCCACAACACGGTTGCTGTA-3′ (Seq. ID No. 48) were used and 15 cycles were practiced. Taq-Gold was used as polymerase. The program of PCR thermal cycler for each clock gene was as follows: degeneration for 9 min at 95° C. only for the first time; followed by repetitive cycles each consisting of thermal degeneration for 15 sec at 94° C., annealing for 30 sec at 55° C. and extension for 30 sec at 72° C.; and finally the extension reaction for 7 min at 72° C.

[0084]FIG. 11 shows the results of the above. It was confirmed by the result that all four kinds of transcripts that were expressed in the chick pineal cells displayed daily fluctuations in abundance with diverged phases and amplitudes in LD cycles and under DD condition. The fluctuation profiles in vivo in Example 3 (FIG. 10) are very similar to those in vitro in Example 4 (FIG. 11), where the cPer2 mRNA levels had a peak early in the morning and a trough early at night. This result was similar to the fluctuation profile of mPerl in the mouse SCN (Cell 90, 1003-1011, 1997, Nature 389, 512-516, 1997). A high level expression of cPer2 sustained at the early light phase (Zeitgeber time (ZT) 2-6) under LD condition, as compared with a rapid decline in cPer2 expression at circadian time (CT) 2-6 under DD condition, indicated that the pineal photoreception plays a role in keeping the high level expression of cPer2 in the morning. The mRNA levels of cBmall and cBmal2 also exhibited clear oscillations and their phases were opposite to that of cPer2 (FIG. 11). Peak time in the cBmal2 mRNA level was delayed by about 4 hr compared to that in the in vitro cBmall mRNA level. This tendency was also observed in the in vivo fluctuation profile. In contrast, the cClock mRNA level showed a relatively low amplitude with a broad peak at ZT 10-18 or CT 10-18, and the peak seems to cover the peaks in expression levels of the two Bmal genes. A similar oscillation of cClock mRNA is observed in the chicken retina (Mol. Brain Res. 70, 253-263, 1999).

EXAMPLE 5 Expression of the Mouse Clock Genes in the Suprachiasmatic Nuclei

[0085] mRNA levels of mBmal2 and known clock genes (mPer2, mClock and mBmall) of the mouse suprachiasmatic nuclei under LD cycles were studied as follows. 5-week-old male C57BL/6 mice were subjected to LD cycles at 23° C.±1° C. (about 200 lux of bright cycle under a fluorescent lamp) and bred with free access to feed and water. 3 weeks thereafter, the mice were decapitated and the brains were rapidly isolated, frozen, and sectioned into thin strips with 700 pm thickness. Small tissue sections including SCN on both sides were taken out from the sections by using a 20-gauge needle, and the mRNA expression levels in mBmal2, mPer2, mClock, mBmall, etc. in the tissue sections were quantified by a quantitative RT-PCR. Three independent RNA samples prepared from six mice (n=3) were respectively quantified and each signal intensity thus obtained was normalized to the signals for mGAPDH and the mean of the three values (mean±SEM) were calculated. p values in FIG. 12 were determined by using Student's t test.

[0086] The above-mentioned primers and number of PCR cycle were determined to give linear relationships between the amounts of the template cDNA and amplification products. For mBmal2, mBMAL2-F2 primer; 5′-TGGTTGGATGCGAAAGAGG-3′ (Seq. ID No. 49) and mBMAL2-R4 primer; 5′-AGGTTTCTCTCTTGGTGAACC-3′ (Seq. ID No. 50) were used and 28 cycles were practiced. For mBmal1 (GenBank accession no. AB012600), rmBmall-F1 primer; 5′-TGGTACCAACATGCAATGC-3′ (Seq. ID No. 51) and rmBmal1-R1 primer; 5′-AGTGTCCGAGGAAGATAGCTG-3′ (Seq. ID No. 52) are used and 28 cycles were practiced. For mPer2 (GenBank accession no. AB016532), rmPer2-Fl primer; 5′-GCTCACTGCCAGAACTATCTCC-3′ (Seq. ID No. 53) and rmPer2-R1 primer; 5′-CCTCTAGCTGAAGCAGGTTAAG-3′ (Seq. ID No. 54) are used and 30 cycles were practiced. For mClock (GenBank accession no. AB019258), rmClock-Fl primer; 5′-CAAGGTCAGCAACTTGTGACC-3′ (Seq. ID No. 55) and rmClock-R1 primer; 5′-AGGATGAGCTGTGTCGAAGG-3′ (Seq. ID No. 56) were used and 28 cycles were practiced. For mGAPDH (GenBank accession no. X02231), rmGAPDH-F1 primer; 5′-CATCACCATCTTCCAGGAGC-3′ (Seq. ID No. 57) and rmGAPDH-Rl primer; 5′-ATTGAGAGCAATGCCAGCC-3′ (Seq. ID No. 58) were used and 21 cycles were practiced. Programming for the PCR thermal cycler for each clock gene was carried out under the condition described in Example 4.

[0087] The results of the above are shown in FIG. 12. In these results, the mPer2 mRNA level displayed daily fluctuations in abundance in the SCN region (FIG. 12A) as are reported in the literatures (Genes Cell 3, 167-176, 1998, Science 288, 1013-1019, 2000). Besides, the mBmall mRNA level showeda faint oscillation in almost antipahse to mPer2 which is in LD cycles (FIG. 12C). On the contrary, mRNA level of mBmal2 was almost constant all day long which was similar to the case of mClock (FIG. 12B, D), suggesting the difference in transcriptional regulation between mBmall and mBmal2 genes.

EXAMPLE 6 Changes in the Photo-Dependency of mRNA Levels in cPer2, cBmal1 and cBmal2 in the Chick Pineal Glands

[0088] Since the expression level of cBmal1/2. in the early morning was low (FIG. 12), a possible light-dependent down-regulation of cBmal1/2 transcriptions was tested. Chicks were exposed to light for a time period when both cBmal1/2 expression levels were high in the dark (CT14-CT15), as is seen from the results of Example 4, and changes in mRNA levels were evaluated at CT15.5 and CT17. One-day-old chicks were entrained to LD cycle for a week and then placed in DD condition. The chick pineal glands that were exposed to a 1-hr light-pulse (350 lux) (CT14-CT15) on the first day of DD condition (FIG. 13A, below) and the chick pineal glands without exposure to light-pulse (FIG. 13A, top) were respectively isolated at CT15.5 or CT17 and the total RNA (8 μg) obtained from each of the pineal glands were respectively separated by an agarose gel electrophoresis and blotted onto a nitrocellulose membrane.

[0089] Each blotting membrane as aforementioned was cut into two pieces and one (containing RNA longer than 2.4 Kb) was hybridized with a cBmall, cBmal2 or cPer2 probe and another with a histone H4 probe. Then the signals for cBmall (FIG. 13B), cBmal2 (FIG. 13C), cPer2 (FIG. 13D) and histone H4 were quantified by MacBAS software (FUJI PHOTO FILM) and the intensity of all the signals were normalized to those for histone H4. The mean value of each gene at CT14 was set to 1 and the mRNA levels were determined. The values were determined from triplicate experiments practiced in a similar way as in the above and shown as mean±SEM. FIG. 13 shows the results. In FIG. 13, “an asterisk” and “double asterisks” mean p <0.05 and p<0.02, respectively. p values were determined using Student's t test. These results demonstrate that mRNA levels of cBmall and cBmal2 observed in the pineal glands of chicks exposed to light at CT15.5 were substantially lower than those of the control animals without exposure to light. On the contrary, the light-induced cPer2 expression was confirmed at CT17, two hours after the exposure to light, as was observed for mPerl and mPer2 in the SCN of the mice exposed to light (Cell 91, 1055-1064, 1997, Neuron 19, 1261-1269, 1997, Genes Cells 3, 167-176, 1998).

EXAMPLE 7 Functional Property of cBMAL2; Pull-Down Assay)

[0090] A close kinship between BMAL1 and BMAL2 among ARNT- (aryl hydrocarbon receptor nuclear translocator) related proteins (FIG. 5) seems to indicate their functional similarity. Therefore, relationships among cBMAL1, cBMAL2 and cCLOCK were tested by a glutathione-S-transferase (GST) pull-down assay using three kinds of bacterially expressed GST-fusion proteins [GST-cCLOCKΔ (a fusion of GST and Met¹-Ser⁴⁶⁶cCLOCK truncated at the carboxy-terminal region), GST-cBMAL1 and GST-cBMAL2], together with [³⁵S]-labeled cBMAL1Δ (Met¹-Ser⁴⁴⁹) or [³⁵S]-labeled cBMAL2Δ(Met¹-Leu⁴⁵⁸) that were transcribed and translated in vitro. Because GST-cCLOCK (a fusion protein composed of GST and the full-length cCLOCK) was not solubilized by 2% Triton X-100, GST- cCLOCKA mentioned above was used instead.

[0091] A DNA fragment encoding GST-cCLOCKA, GST-BMAL1, GST-BMAL2 or GST, mentioned above, was introduced into a pGEX5X-1 expression vector and expressed in BL21 E. coli strain. Each E. coli was subjected to lysis in buffer A [10 mM Na-phosphate (pH 7.9), 140 mM NaCl, 1 mM MgCl₂, 10 mM EDTA, 5 mM 2-mercaptoethanol, 2 mM PMSF and one tablet of Complete EDTA-free protease inhibitor (Roche Diagnostics) per 50 mL], then each of solubilized fusion proteins or GST was purified by glutathione-Sepharose column (Amersham Pharmacia Biotech). On the other hand, [³⁵S]-labeled cBMAL1Δ (Met¹-Ser⁴⁴⁹) and [³⁵S]-labeled cBMAL2Δ (Met¹-Leu⁴⁵⁸) mentioned above were prepared by the in vitro transcription and translation of an expression plasmid containing CDNA fragment of cBMAL1Δ (Met¹-Ser⁴⁴⁹) or cBMAL2Δ (Met¹-Leu⁴⁵⁸) in the presence of [³⁵S] methionine and with the aid of TNT-T7 Quick Coupled Transcription/Translation System (Promega). [³⁵S]-labeled luciferase as a control was similarly transcribed and translated in vitro.

[0092] 8 μL each of the [³⁵S]-labeled protein (cBMAL1Δ, cBMAL2Δ or luciferase protein) solutions was mixed with 40 μL of glutathione-sepharose beads, to which GST-cCLOCKA (0.1 μg), GST-cBMAL1 (1.1 μg), GST-cBMAL2 (3.3 μg) or GST (5.6 pg) had been bound. Then the mixtures were incubated in 140 μL of buffer B [20 mM Hepes-NaOH (pH 7.9), 20% (w/v) glycerol, 15 mM KC1, 0.2% Triton X-100, 2.5% skim milk, one tablet of Complete EDTA-free protease inhibitor per 50 mL] on ice for 1 hr with gentle rotation. After the incubation the mixtures were washed four times with buffer C [10 mM Tris-HCl (pH 7.5), 0.2% Triton X-100, 150 mM NaCl, 2 mM EDTA, 1 mM PMSF, one tablet of Complete EDTA-free protease inhibitor] and were separated by a SDS- polyacrylamide (10%) gel electrophoresis, then the gel was analyzed for autoradiograph by using a FLA2000 bioimage analyzer (FUJI PHOTO FILM).

[0093] The results of the above are shown in FIG. 14. Lanes 16-18 is the results of electrophoresis for [³⁵S]-labeled cBMAL1Δ, cBMAL2Δ or luciferase (2.5% each of the inputs). A faint signal observed in lane 17 (the upper band) is due to the migration of luciferase from lane 18. These results revealed that GST-cCLOCKΔ specifically bound not only with cBMAL1Δ but also with cBMAL2Δ in vitro (FIG. 14, lanes 1, 2). Interestingly, GST-cBMAL2 bound with both cBMAL proteins (FIG. 14, lanes 4, 5), and GST-cBMALl also showed similar binding profiles (FIG. 14, lanes 7, 8), indicating potential activity of cBMAL proteins to form a homodimer as well as a cBMAL1-cBMAL2 heterodimer. It was also demonstrated that a CBMAL protein deficient in the C-terminal bound more efficiently with a GST-fusion protein than with a full-length CBMAL protein.

EXAMPLE 8 an Electrophoretic Mobility Shift Assay Using a cPer2 E-box-Containing Probe

[0094] A binding of cBMAL1-cCLOCK or cBMAL2-cCLOCK to the E-box sequence was examined by an electrophoretic mobility shift assay (EMSA) in which an E-box (CACGTG)-containing sequence present in a promoter region of cPer2 gene was used as a probe. For preparation of the probe, oligonucleotides [cP2E1-S: 5′-GTGTCACACGTGAGGCTTA-3′ (Seq. ID No. 59) and cP2E1-AS: 5′-TAAGCCTCACGTGTGACAC-3′ (Seq. ID No. 60)] were synthesized that correspond to the E-box sequence and its flanking sequences within a putative promoter/enhancer region of cPer2 gene. These oligonucleotides synthesized were annealed together and subcloned into a pCR2.1 vector using TOPO-TA cloning kit (Invitrogen, Calif.), from which a 39 bp fragment was excised with a restriction enzyme EcoRI and used. The above-mentioned cBMAL1, cBMAL2 and cCLOCK were prepared by being transcribed and translated in vitro from an expression plasmid containing the cDNA of cBmal1, cBmal2 or cClock with the aid of TNT-T7 Quick Coupled Transcription/Translation System (Promega). A pcDNA3.1/V5/His empty vector, an expression vector, alone was transcribed and translated similarly as in the above and used as a control.

[0095] 5 μL each of the protein mixtures thus obtained (BMAL1+BMAL2, BMAL1+CLOCK, BMAL2+CLOCK) was added with 32 μL of buffer [25 mM Hepes-KOH (pH 7.6), 100 mM KCl, 0.1 mM EDTA, 10% (v/v) glycerol, 7.5 mM MgCl₂, 1 mM DTT and 1 μg denatured salmon sperm DNA] containing a ³²P-labeled probe (33 fmoles, 1.3×10⁵ cpm) andwas incubated for 20 min at 23° C. After the incubation, each mixture was separated by a 6% polyacrylamide gel electrophoresis and analyzed similarly as in Example 7 using a FLA2000 bioimage analyzer (FUJI PHOTO FILM). FIG. 15 shows the results. In FIG. 15, lane 1 is the result of the labeled probe alone, lanes 2-5 are the results of the reactions between each translation product (control, BMAL1, BMAL2 or CLOCK) and the labeled probe. In the figure, the asterisk denotes the position of the free probe, closed arrowheads represent specific complexes with the bHLH-PAS proteins, and open arrowheads indicate background. It was confirmed from these results that in the presence of cCLOCK, cBMAL2 and cBMAL1 had respectively formed two or three complexes (closed arrowheads in lanes 7 and 8 in FIG. 15). It is unlikely that these complexes represent homodimers of any of the PAS proteins examined (cCLOCK, cBMAL1 or cBMAL2), because no specific bands were observed when cCLOCK, cBMALl or cBMAL2 alone was reacted with the probe (lanes 3-6 in FIG. 15). These results suggest that the cPer2 E-box is one of the in vivo targets of cCLOCK-cBMAL1/2 heteromer.

EXAMPLE 9 Transcriptional Regulation by cBMAL1, cBMAL2 and cCLOCK in 293EBNA Cells

[0096] Abilities for the transcriptional activation and suppression of cBMAL1, cBMAL2 and cCLOCK were tested with the mPer2 E-box or the mPerl promoter as a role model in the feed-back-loop and the vasopressin gene E-box as a role model in output pathways. Human embryonic kidney 293EBNA cells (Invitrogen) were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum (Life Technologies) which cultured cells were then plated at 3+10⁵ cells per well on six-well plates and transfected by using a total of 1.0 μg of various expression plasmids [an expression vector, plasmids containing reporter genes, 0.25 ng of Renilla luciferase reporter (pRL-CMV; Promega), and plasmids containing cDNAof each clock gene (cBmal1, cBmal2, cClock) with the amount indicated in FIG. 16] together with Lipofectamine plus (Life Technologies).

[0097] As for the expression vector mentioned above, pcDNA3.1/V5/Hisemptyvector(Invitrogen)wasused. As forthe reporter genes mentioned above, 25 ng of the firefly luciferase reporter (cPer2 E-box-luc; a derivative of pGL3-Promoter; Promega) containing mPer2 E-box, 25 ng of cPer2 mut.E-box-luc, 50 ng of the firefly luciferase reporter containing mPerl promoter (mPerl-luc; a derivative of pGL3-Basic; Promega) 25 ng of the firefly luciferase reporter containing the mouse vasopressin E-box (AVP E-box-luc; a derivative of pGL3-Promoter; Promega), 25 ng of AVP mut. E-box-luc, or 25 ng of TRE-luc were used. Two days after the transfection, cell extracts were subjected to dual-luciferase assays by luminometry (Promega) according to the manufacturer's protocol. For each extract, the firefly luciferase activity was normalized by the Renilla luciferase activity and the mean value (means±SEM) was determined from the values of three independent culture extracts.

[0098] The aforementioned plasmids containing reporter genes were prepared as follows. The E-box sequence, CACGTG and its flanking sequences within the promoter/enhancer region of cPer2 gene were linked in tandem (5′-GTGTCACACGTGAGGCTTAGTGTCACACGTGAGGCTTAGTGTCACACGTGAGGCTTA-3′), which was then inserted into a luciferase reporter containing SV-40 (pGL3-Promoter, Promega) and thus the cPer2 E-box-luc was constructed. The cPer2 mut.E-box as a reporter plasmid in the control experiment was constructed by mutating the E-box sequences into GGACCT in a similar way as previously reported (Cell 96, 57-68, 1999). mperl-luc was constructed as follows; a 2.2 Kb upstream fragment of mPerl was amplified by PCR using the DNA templates from the mouse genome [sense primer 3; 5′-TCGAGCTCTTTGGTACCTGGCCAGCAACC-3′ (Seq. ID No. 61) and anti-senseprimer3; 5′-TCACGACACCTGGCCGTTCGAGG-3′ (Seq. IDNo. 62)] and LA-Taq polymerase, base sequences for the six clones individually obtained by PCR were determined, and then one clone without PCR error among the six clones was linked to a luciferase reporter (pGL3-Basic, Promega). AVP E-box-luc was constructed by linking E-box sequence (CACGTG) in the promoter/enhancer region in the mouse vasopressin gene and its flanking sequences, and then by inserting the resulting sequence (5′-TCAGGCCCACGTGTCCCA-3′) into the luciferase reporter containing SV-40 promoter (pGL3-Promoter, Promega). Further, the AVP mut. E-box-luc (a reporter with a E-box mutation) which is a reporter plasmid for the control experiment was prepared in a way previously described (Cell 96, 57-68, 1999). TRE-luc was prepared as follows; the phorbol ester-responsive element (TRE) and its flanking sequences within human collagenase gene were linked in tandem [5′-CGGCTGACTCATCAAGCTGACTCATCAAGCTGACTCATCAA-3′ (Seq. ID No. 63)],whichwastheninsertedintotheBglIIsiteinaluciferase reporter in which a BglII-HindIII fragment of pRL-TK vector (Promega) was ligated to a pGL3-Basic vector (pGL3-TK-promoter vector).

[0099] These results are shown in FIG. 16. The results show that cCLOCK binds to not only cBMAL1 but cBMAL2 and promotes the transactivation which is medidted by the cPer2 E-box (FIG. 16A). Similar results were obtained by using a 2.2-kb mPer1 promoter harboring three E-box sequences (CACGTG) (FIG. 16B). Interestingly, the transactivation elicited by cBMAL2-cCLOCK showed a clear peak when a relatively low dose (20 ng) of a cBmal2 expression plasmid and cClock plasmid (250 ng) were coexpressed, and a higher dose than the above of cBmal2 plasmid suppressed the transactivation in FIG. 16B (see the left of the figure) and FIG. 16C (see 10th-16th bars from the left of the figure). cBmall, however, seems to have no such inhibitory effect. Endogenous transactivation neither from the TPA-responsive element (TRE, FIG. 16B) nor from the SV40-promoter was suppressed by application of a high dose (160 ng) of cBmal2, which fact suggests that the suppression is due to the specific effect on E-box or E-box-binding component(s).

[0100] Since cBmal1 and cBmal2 had the slightly shifted expression profiles as can be seen in FIGS. 10 and 11, a cooperative effect of cBMAL1 and cBMAL2 on the transcriptional regulation was tested. In the case of a vasopressin gene E-box as a reporter (FIG. 16C), a low level expression (10ng) of cBmal2 notably enhanced cBMAL1-cCLOCK transactivation (see 17th-23rd bars from the left in FIG. 16C). A similar or more pronounced cooperative effect was observed with a low dose of cBmal1 plasmid (10 ng) for cBMAL2-cCLOCK transactivation (see 24th-30th bars from the left in FIG. 16C). Besides, the cooperative activation was considerably suppressed by the application of larger amounts of cBmal2 (80-160 ng) or cBmall (40-160 ng). Similar results were also observed in the cases when a cPer2 E-box or a mPerl promoter was used, albeit with less degrees (FIG. 16B).

EXAMPLE 10 Effect of cPER2 on Transactivation Mediated by E-box Sequences

[0101] Next, whether cPER2 negatively acted on the transactivation elicited by the transactivator cBMAL-cCLOCK was examined. The experiment described in this Example 10 was performed in a similar way as in Example 9 except that plasmids containing a cPer2 cDNA were transfected, with the amounts shown in FIGS. 17A and 17B, to the expression plasmids which were to betransfectedtothehumanembryonickidney293EBNAcells. The results are shown in FIGS. 17A and 17B. The results show that coexpression of cPer2 plasmid (250 ng) in 293EBNA cells inhibited the cBMAL2-cCLOCK-dependent transactivation mediated by cPer2 E-box, and the degree of the inhibitory effect was stronger than that on cBMALl-cCLOCK-dependent transactivation under the same conditions (FIG. 17A). Similar tendency was also observed in the case of cBMAL-cCLOCK-dependent transactivation mediated by the vasopressin E-box (FIG. 17B), and the higher degree of inhibitory effect was observed with the increase in the cPER2 amount.

[0102] Then, intrinsic properties of the cPer2 E-box mediated transactivation were studied in the cultured chick pineal cells. The pineal cells prepared from one-day-old chicks were plated at 4×10⁵ cells per well on 24-well plates and cultured under LD cycle. At ZT9 on Day 3 of the culture, the pineal cells were transfected with 500 ng of either the aforementioned cPer2 expression plasmid or pcDNA3.1/V5/His (control), 250 ng of either the cPer2 E-box-luc or the cPer2 mut.E-box-luc, and 5 ng of pRL-CMV (Promega) by using Lipofectamine plus. At ZT6 on the next day of the transfection, the cell extracts were subjected to a dual-luciferase assay and the results are shown in FIG. 17C. The results demonstrated that the endogenous transactivation mediated by cPer2 E-box was markedly decreased as a result of mutating the E-box sequence and that the inhibitory effect on transactivation induced by forced expression of cPER2 was also E-box-dependent. These facts suggest that the chicken pineal cells express a positive factor acting on the cPer2 E-box and that this factor exhibits an effect on the negative regulation by cPER2.

EXAMPLE 11 Ablation of Melatonin Rhythm by the Overexpression of cBMAL1 and cBMAL2

[0103] cBMAL1 or cBMAL2 was overexpressed in the cultured chick pineal cells and its effect on the melatonin rhythm was examined to evaluate the roles of the two PAS proteins in maintenance of the rhythmicity. The chick pineal cells were cultured in 24-well cloning plates (Greiner Labortechnik, Frickenhausen, Germany) for 2 days and transfected with 500 ng of either cBMAL1 or cBMAL2 expression plasmid mentioned above or pcDNA3. 1/V5/His (control) by using a combination of Lipofectamine plus (Life Technologies) and Genefector (VennNova LLc, FL). 2 days after the transfection, the cells were subjected to a 4-day culture in the media containing 200 mg/L G418 (Life Technologies) to select the transfected cells and the cells selected were further cultured in the media containing 50 mg/L G418. The culture media were collected every 4 hours to quantify the released melatonin by the previously described method (Neurosci 20, 986-991, 2000). FIG. 18 shows the results. Four data in each panel are the results obtained from the individual cultures where each value was determined by setting the average of melatonin production levels during the LD cycles to 1. The bar at the bottom of FIG. 18 represents lighting conditions.

[0104] A slight phase-delaying was observed upon studying the melatonin rhythm in the pineal gland of each cell. This change was also observed in the untransfected pineal cells, and such clock oscillation was also observed after culturing control cells (FIG. 9A) and cells overexpressing proteins unrelated to clock proteins such as a ml or m2 acetylcholine receptor, under DD condition for several days. In contrast to these control cells, cBMAL1- or cBMAL2-overexpressing cells displayed only a single oscillation in melatonin production under DD condition, which was thereafter kept at a constant level (FIGS. 18B and 18C). Under the LD cycles, daily melatonin fluctuations in cBMALl- or cBMAL2-overexpressing cells were quite similar to those of control cells, indicating that cellular mechanisms for light-dependent melatonin production were stably maintained by the overexpressed cBMAL proteins. In spite of this, the ablation of rhythm under DD condition strongly suggests that cBMAL1 and cBMAL2 are both indispensable factors for rhythmic oscillation.

INDUSTRIAL APPLICABILOTY

[0105] The present invention makes it possible to provide novel clock proteins having the novel BMAL2 activity crucial for the clock oscillation mechanism including photic-input pathway and output pathway, and the gene DNAs encoding the proteins. Further, with the use of these proteins and the gene DNAs, substances useful for prevention and therapy of the circadian rhythm sleep disorders or the like including delayed sleep phase syndrome, non-24-hour sleep-wake syndrome, advanced sleep phase syndrome, time zone change syndrome, shift work sleep disorder, etc. can be screened, in addition to which a molecular mechanism of the circadian oscillation system can also be elucidated. Still further, the proteins of the present invention having the BMAL2 activity have functions both for promoting and suppressing transcription and are thought to be involved in diverse biological functions by binding with partners other than CLOCK. The proteins are therefore expected to be applied to specifically inhibit a group of functions in the transcriptional regulatory regions including that of period genes by gene-introduction of BMAL2 or the BMAL2-dominant negative mutants in an excessive amount from the outside.

1 63 1 1930 DNA Homo sapiens CDS (19)..(1926) 1 gaccaagtgg ctcctgcg atg gcg gcg gaa gag gag gct gcg gcg gga ggt 51 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly 1 5 10 aaa gtg ttg aga gag gag aac cag tgc att gct cct gtg gtt tcc agc 99 Lys Val Leu Arg Glu Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser 15 20 25 cgc gtg agt cca ggg aca aga cca aca gct atg ggg tct ttc agc tca 147 Arg Val Ser Pro Gly Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser 30 35 40 cac atg aca gag ttt cca cga aaa cgc aaa gga agt gat tca gac cca 195 His Met Thr Glu Phe Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro 45 50 55 tcc cag tca gga atc atg aca gaa aaa gtg gtg gaa aag ctt tct cag 243 Ser Gln Ser Gly Ile Met Thr Glu Lys Val Val Glu Lys Leu Ser Gln 60 65 70 75 aat ccc ctt acc tat ctt ctt tca aca agg ata gaa ata tca gcc tcc 291 Asn Pro Leu Thr Tyr Leu Leu Ser Thr Arg Ile Glu Ile Ser Ala Ser 80 85 90 agt ggc agc aga gtg gaa gat ggt gaa cac caa gtt aaa atg aag gcc 339 Ser Gly Ser Arg Val Glu Asp Gly Glu His Gln Val Lys Met Lys Ala 95 100 105 ttc aga gaa gct cat agc caa act gaa aag cgg agg aga gat aaa atg 387 Phe Arg Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met 110 115 120 aat aac ctg att gaa gaa ctg tct gca atg atc cct cag tgc aac ccc 435 Asn Asn Leu Ile Glu Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro 125 130 135 atg gcg cgt aaa ctg gac aaa ctt aca gtt tta aga atg gct gtt caa 483 Met Ala Arg Lys Leu Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln 140 145 150 155 cac ttg aga tct tta aaa ggc ttg aca aat tct tat gtg gga agt aat 531 His Leu Arg Ser Leu Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn 160 165 170 tat aga cca tca ttt ctt cag gat aat gag ctc aga cat tta atc ctt 579 Tyr Arg Pro Ser Phe Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu 175 180 185 aag act gca gaa ggc ttc tta ttt gtg gtt gga tgt gaa aga gga aaa 627 Lys Thr Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys 190 195 200 att ctc ttc gtt tct aag tca gtc tcc aaa ata ctt aat tat gat cag 675 Ile Leu Phe Val Ser Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln 205 210 215 gct agt ttg act gga caa agc tta ttt gac ttc tta cat cca aaa gat 723 Ala Ser Leu Thr Gly Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp 220 225 230 235 gtt gcc aaa gta aag gaa caa ctt tct tct ttt gat att tca cca aga 771 Val Ala Lys Val Lys Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg 240 245 250 gaa aag cta ata gat gcc aaa act ggt ttg caa gtt cac agt aat ctc 819 Glu Lys Leu Ile Asp Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu 255 260 265 cac gct gga agg aca cgt gtg tat tct ggc tca aga cga tct ttt ttc 867 His Ala Gly Arg Thr Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe 270 275 280 tgt cgg ata aag agt tgt aaa atc tct gtc aaa gaa gag cat gga tgc 915 Cys Arg Ile Lys Ser Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys 285 290 295 tta ccc aac tca aag aag aaa gag cac aga aaa ttc tat act atc cat 963 Leu Pro Asn Ser Lys Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His 300 305 310 315 tgc act ggt tac ttg aga agc tgg cct cca aat att gtt gga atg gaa 1011 Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu 320 325 330 gaa gaa agg aac agt aag aaa gac aac agt aat ttt acc tgc ctt gtg 1059 Glu Glu Arg Asn Ser Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val 335 340 345 gcc att gga aga tta cag cca tat att gtt cca cag aac agt gga gag 1107 Ala Ile Gly Arg Leu Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu 350 355 360 att aat gtg aaa cca act gaa ttt ata acc cgg ttt gca gtg aat gga 1155 Ile Asn Val Lys Pro Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly 365 370 375 aaa ttt gtc tat gta gat caa agg gca aca gcg att tta gga tat ctg 1203 Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu 380 385 390 395 cct cag gaa ctt ttg gga act tct tgt tat gaa tat ttt cat caa gat 1251 Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp 400 405 410 gac cac aat aat ttg act gac aag cac aaa gca gtt cta cag agt aag 1299 Asp His Asn Asn Leu Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys 415 420 425 gag aaa ata ctt aca gat tcc tac aaa ttc aga gca aaa gat ggc tct 1347 Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser 430 435 440 ttt gta act tta aaa agc caa tgg ttt agt ttc aca aat cct tgg aca 1395 Phe Val Thr Leu Lys Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr 445 450 455 aaa gaa ctg gaa tat att gta tct gtc aac act tta gtt ttg gga cat 1443 Lys Glu Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly His 460 465 470 475 agt gag cct gga gaa gca tca ttt tta cct tgt agc tct caa tca tca 1491 Ser Glu Pro Gly Glu Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser 480 485 490 gaa gaa tcc tct aga cag tcc tgt atg agt gta cct gga atg tct act 1539 Glu Glu Ser Ser Arg Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr 495 500 505 gga aca gta ctt ggt gct ggt agt att gga aca gat att gca aat gaa 1587 Gly Thr Val Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu 510 515 520 att ctg gat tta cag agg tta cag tct tct tca tac ctt gat gat tcg 1635 Ile Leu Asp Leu Gln Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser 525 530 535 agt cca aca ggt tta atg aaa gat act cat act gta aac tgc agg agt 1683 Ser Pro Thr Gly Leu Met Lys Asp Thr His Thr Val Asn Cys Arg Ser 540 545 550 555 atg tca aat aag gag ttg ttt cca cca agt cct tct gaa atg ggg gag 1731 Met Ser Asn Lys Glu Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu 560 565 570 cta gag gct acc agg caa aac cag agt act gtt gct gtc cac agc cat 1779 Leu Glu Ala Thr Arg Gln Asn Gln Ser Thr Val Ala Val His Ser His 575 580 585 gag cca ctc ctc agt gat ggt gca cag ttg gat ttc gat gcc cta tgt 1827 Glu Pro Leu Leu Ser Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys 590 595 600 gac aat gat gac aca gcc atg gct gca ttt atg aat tac tta gaa gca 1875 Asp Asn Asp Asp Thr Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala 605 610 615 gag ggg ggc ctg gga gac cct ggg gac ttc agt gac atc cag tgg acc 1923 Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr 620 625 630 635 ctc tagc 1930 Leu 2 636 PRT Homo sapiens 2 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly Lys Val Leu Arg Glu 1 5 10 15 Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser Arg Val Ser Pro Gly 20 25 30 Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser His Met Thr Glu Phe 35 40 45 Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro Ser Gln Ser Gly Ile 50 55 60 Met Thr Glu Lys Val Val Glu Lys Leu Ser Gln Asn Pro Leu Thr Tyr 65 70 75 80 Leu Leu Ser Thr Arg Ile Glu Ile Ser Ala Ser Ser Gly Ser Arg Val 85 90 95 Glu Asp Gly Glu His Gln Val Lys Met Lys Ala Phe Arg Glu Ala His 100 105 110 Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn Asn Leu Ile Glu 115 120 125 Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro Met Ala Arg Lys Leu 130 135 140 Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln His Leu Arg Ser Leu 145 150 155 160 Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn Tyr Arg Pro Ser Phe 165 170 175 Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu Lys Thr Ala Glu Gly 180 185 190 Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys Ile Leu Phe Val Ser 195 200 205 Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln Ala Ser Leu Thr Gly 210 215 220 Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys 225 230 235 240 Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg Glu Lys Leu Ile Asp 245 250 255 Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu His Ala Gly Arg Thr 260 265 270 Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe Cys Arg Ile Lys Ser 275 280 285 Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys Leu Pro Asn Ser Lys 290 295 300 Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His Cys Thr Gly Tyr Leu 305 310 315 320 Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu Glu Glu Arg Asn Ser 325 330 335 Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val Ala Ile Gly Arg Leu 340 345 350 Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu Ile Asn Val Lys Pro 355 360 365 Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly Lys Phe Val Tyr Val 370 375 380 Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu 385 390 395 400 Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Asn Asn Leu 405 410 415 Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr 420 425 430 Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser Phe Val Thr Leu Lys 435 440 445 Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Glu Leu Glu Tyr 450 455 460 Ile Val Ser Val Asn Thr Leu Val Leu Gly His Ser Glu Pro Gly Glu 465 470 475 480 Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser Glu Glu Ser Ser Arg 485 490 495 Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr Gly Thr Val Leu Gly 500 505 510 Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Ile Leu Asp Leu Gln 515 520 525 Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser Ser Pro Thr Gly Leu 530 535 540 Met Lys Asp Thr His Thr Val Asn Cys Arg Ser Met Ser Asn Lys Glu 545 550 555 560 Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu Leu Glu Ala Thr Arg 565 570 575 Gln Asn Gln Ser Thr Val Ala Val His Ser His Glu Pro Leu Leu Ser 580 585 590 Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys Asp Asn Asp Asp Thr 595 600 605 Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly 610 615 620 Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr Leu 625 630 635 3 1888 DNA Homo sapiens CDS (19)..(1884) 3 gaccaagtgg ctcctgcg atg gcg gcg gaa gag gag gct gcg gcg gga ggt 51 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly 1 5 10 aaa gtg ttg aga gag gag aac cag tgc att gct cct gtg gtt tcc agc 99 Lys Val Leu Arg Glu Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser 15 20 25 cgc gtg agt cca ggg aca aga cca aca gct atg ggg tct ttc agc tca 147 Arg Val Ser Pro Gly Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser 30 35 40 cac atg aca gag ttt cca cga aaa cgc aaa gga agt gat tca gac cca 195 His Met Thr Glu Phe Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro 45 50 55 tcc cag tca gga atc atg aca gaa aaa gtg gtg gaa aag ctt tct cag 243 Ser Gln Ser Gly Ile Met Thr Glu Lys Val Val Glu Lys Leu Ser Gln 60 65 70 75 aat ccc ctt acc tat ctt ctt tca aca agg ata gaa ata tca gcc tcc 291 Asn Pro Leu Thr Tyr Leu Leu Ser Thr Arg Ile Glu Ile Ser Ala Ser 80 85 90 agt ggc agc aga gaa gct cat agc caa act gaa aag cgg agg aga gat 339 Ser Gly Ser Arg Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp 95 100 105 aaa atg aat aac ctg att gaa gaa ctg tct gca atg atc cct cag tgc 387 Lys Met Asn Asn Leu Ile Glu Glu Leu Ser Ala Met Ile Pro Gln Cys 110 115 120 aac ccc atg gcg cgt aaa ctg gac aaa ctt aca gtt tta aga atg gct 435 Asn Pro Met Ala Arg Lys Leu Asp Lys Leu Thr Val Leu Arg Met Ala 125 130 135 gtt caa cac ttg aga tct tta aaa ggc ttg aca aat tct tat gtg gga 483 Val Gln His Leu Arg Ser Leu Lys Gly Leu Thr Asn Ser Tyr Val Gly 140 145 150 155 agt aat tat aga cca tca ttt ctt cag gat aat gag ctc aga cat tta 531 Ser Asn Tyr Arg Pro Ser Phe Leu Gln Asp Asn Glu Leu Arg His Leu 160 165 170 atc ctt aag act gca gaa ggc ttc tta ttt gtg gtt gga tgt gaa aga 579 Ile Leu Lys Thr Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg 175 180 185 gga aaa att ctc ttc gtt tct aag tca gtc tcc aaa ata ctt aat tat 627 Gly Lys Ile Leu Phe Val Ser Lys Ser Val Ser Lys Ile Leu Asn Tyr 190 195 200 gat cag gct agt ttg act gga caa agc tta ttt gac ttc tta cat cca 675 Asp Gln Ala Ser Leu Thr Gly Gln Ser Leu Phe Asp Phe Leu His Pro 205 210 215 aaa gat gtt gcc aaa gta aag gaa caa ctt tct tct ttt gat att tca 723 Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser Ser Phe Asp Ile Ser 220 225 230 235 cca aga gaa aag cta ata gat gcc aaa act ggt ttg caa gtt cac agt 771 Pro Arg Glu Lys Leu Ile Asp Ala Lys Thr Gly Leu Gln Val His Ser 240 245 250 aat ctc cac gct gga agg aca cgt gtg tat tct ggc tca aga cga tct 819 Asn Leu His Ala Gly Arg Thr Arg Val Tyr Ser Gly Ser Arg Arg Ser 255 260 265 ttt ttc tgt cgg ata aag agt tgt aaa atc tct gtc aaa gaa gag cat 867 Phe Phe Cys Arg Ile Lys Ser Cys Lys Ile Ser Val Lys Glu Glu His 270 275 280 gga tgc tta ccc aac tca aag aag aaa gag cac aga aaa ttc tat act 915 Gly Cys Leu Pro Asn Ser Lys Lys Lys Glu His Arg Lys Phe Tyr Thr 285 290 295 atc cat tgc act ggt tac ttg aga agc tgg cct cca aat att gtt gga 963 Ile His Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro Asn Ile Val Gly 300 305 310 315 atg gaa gaa gaa agg aac agt aag aaa gac aac agt aat ttt acc tgc 1011 Met Glu Glu Glu Arg Asn Ser Lys Lys Asp Asn Ser Asn Phe Thr Cys 320 325 330 ctt gtg gcc att gga aga tta cag cca tat att gtt cca cag aac agt 1059 Leu Val Ala Ile Gly Arg Leu Gln Pro Tyr Ile Val Pro Gln Asn Ser 335 340 345 gga gag att aat gtg aaa cca act gaa ttt ata acc cgg ttt gca gtg 1107 Gly Glu Ile Asn Val Lys Pro Thr Glu Phe Ile Thr Arg Phe Ala Val 350 355 360 aat gga aaa ttt gtc tat gta gat caa agg gca aca gcg att tta gga 1155 Asn Gly Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly 365 370 375 tat ctg cct cag gaa ctt ttg gga act tct tgt tat gaa tat ttt cat 1203 Tyr Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His 380 385 390 395 caa gat gac cac aat aat ttg act gac aag cac aaa gca gtt cta cag 1251 Gln Asp Asp His Asn Asn Leu Thr Asp Lys His Lys Ala Val Leu Gln 400 405 410 agt aag gag aaa ata ctt aca gat tcc tac aaa ttc aga gca aaa gat 1299 Ser Lys Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Ala Lys Asp 415 420 425 ggc tct ttt gta act tta aaa agc caa tgg ttt agt ttc aca aat cct 1347 Gly Ser Phe Val Thr Leu Lys Ser Gln Trp Phe Ser Phe Thr Asn Pro 430 435 440 tgg aca aaa gaa ctg gaa tat att gta tct gtc aac act tta gtt ttg 1395 Trp Thr Lys Glu Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu 445 450 455 gga cat agt gag cct gga gaa gca tca ttt tta cct tgt agc tct caa 1443 Gly His Ser Glu Pro Gly Glu Ala Ser Phe Leu Pro Cys Ser Ser Gln 460 465 470 475 tca tca gaa gaa tcc tct aga cag tcc tgt atg agt gta cct gga atg 1491 Ser Ser Glu Glu Ser Ser Arg Gln Ser Cys Met Ser Val Pro Gly Met 480 485 490 tct act gga aca gta ctt ggt gct ggt agt att gga aca gat att gca 1539 Ser Thr Gly Thr Val Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala 495 500 505 aat gaa att ctg gat tta cag agg tta cag tct tct tca tac ctt gat 1587 Asn Glu Ile Leu Asp Leu Gln Arg Leu Gln Ser Ser Ser Tyr Leu Asp 510 515 520 gat tcg agt cca aca ggt tta atg aaa gat act cat act gta aac tgc 1635 Asp Ser Ser Pro Thr Gly Leu Met Lys Asp Thr His Thr Val Asn Cys 525 530 535 agg agt atg tca aat aag gag ttg ttt cca cca agt cct tct gaa atg 1683 Arg Ser Met Ser Asn Lys Glu Leu Phe Pro Pro Ser Pro Ser Glu Met 540 545 550 555 ggg gag cta gag gct acc agg caa aac cag agt act gtt gct gtc cac 1731 Gly Glu Leu Glu Ala Thr Arg Gln Asn Gln Ser Thr Val Ala Val His 560 565 570 agc cat gag cca ctc ctc agt gat ggt gca cag ttg gat ttc gat gcc 1779 Ser His Glu Pro Leu Leu Ser Asp Gly Ala Gln Leu Asp Phe Asp Ala 575 580 585 cta tgt gac aat gat gac aca gcc atg gct gca ttt atg aat tac tta 1827 Leu Cys Asp Asn Asp Asp Thr Ala Met Ala Ala Phe Met Asn Tyr Leu 590 595 600 gaa gca gag ggg ggc ctg gga gac cct ggg gac ttc agt gac atc cag 1875 Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe Ser Asp Ile Gln 605 610 615 tgg acc ctc tagc 1888 Trp Thr Leu 620 4 622 PRT Homo sapiens 4 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly Lys Val Leu Arg Glu 1 5 10 15 Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser Arg Val Ser Pro Gly 20 25 30 Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser His Met Thr Glu Phe 35 40 45 Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro Ser Gln Ser Gly Ile 50 55 60 Met Thr Glu Lys Val Val Glu Lys Leu Ser Gln Asn Pro Leu Thr Tyr 65 70 75 80 Leu Leu Ser Thr Arg Ile Glu Ile Ser Ala Ser Ser Gly Ser Arg Glu 85 90 95 Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn Asn Leu 100 105 110 Ile Glu Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro Met Ala Arg 115 120 125 Lys Leu Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln His Leu Arg 130 135 140 Ser Leu Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn Tyr Arg Pro 145 150 155 160 Ser Phe Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu Lys Thr Ala 165 170 175 Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys Ile Leu Phe 180 185 190 Val Ser Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln Ala Ser Leu 195 200 205 Thr Gly Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys 210 215 220 Val Lys Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg Glu Lys Leu 225 230 235 240 Ile Asp Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu His Ala Gly 245 250 255 Arg Thr Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe Cys Arg Ile 260 265 270 Lys Ser Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys Leu Pro Asn 275 280 285 Ser Lys Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His Cys Thr Gly 290 295 300 Tyr Leu Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu Glu Glu Arg 305 310 315 320 Asn Ser Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val Ala Ile Gly 325 330 335 Arg Leu Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu Ile Asn Val 340 345 350 Lys Pro Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly Lys Phe Val 355 360 365 Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu 370 375 380 Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Asn 385 390 395 400 Asn Leu Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile 405 410 415 Leu Thr Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser Phe Val Thr 420 425 430 Leu Lys Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Glu Leu 435 440 445 Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly His Ser Glu Pro 450 455 460 Gly Glu Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser Glu Glu Ser 465 470 475 480 Ser Arg Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr Gly Thr Val 485 490 495 Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Ile Leu Asp 500 505 510 Leu Gln Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser Ser Pro Thr 515 520 525 Gly Leu Met Lys Asp Thr His Thr Val Asn Cys Arg Ser Met Ser Asn 530 535 540 Lys Glu Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu Leu Glu Ala 545 550 555 560 Thr Arg Gln Asn Gln Ser Thr Val Ala Val His Ser His Glu Pro Leu 565 570 575 Leu Ser Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys Asp Asn Asp 580 585 590 Asp Thr Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly 595 600 605 Leu Gly Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr Leu 610 615 620 5 1819 DNA Homo sapiens CDS (19)..(1815) 5 gaccaagtgg ctcctgcg atg gcg gcg gaa gag gag gct gcg gcg gga ggt 51 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly 1 5 10 gag gtt gcc ggt ggc gag gcg acg gcc cca ggt aaa gtg ttg aga gag 99 Glu Val Ala Gly Gly Glu Ala Thr Ala Pro Gly Lys Val Leu Arg Glu 15 20 25 gag aac cag tgc att gct cct gtg gtt tcc agc cgc gtg agt cca ggg 147 Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser Arg Val Ser Pro Gly 30 35 40 aca aga cca aca gct atg ggg tct ttc agc tca cac atg aca gag ttt 195 Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser His Met Thr Glu Phe 45 50 55 cca cga aaa cgc aaa gga agt gat tca gac cca tcc caa gaa gct cat 243 Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro Ser Gln Glu Ala His 60 65 70 75 agc caa act gaa aag cgg agg aga gat aaa atg aat aac ctg att gaa 291 Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn Asn Leu Ile Glu 80 85 90 gaa ctg tct gca atg atc cct cag tgc aac ccc atg gcg cgt aaa ctg 339 Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro Met Ala Arg Lys Leu 95 100 105 gac aaa ctt aca gtt tta aga atg gct gtt caa cac ttg aga tct tta 387 Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln His Leu Arg Ser Leu 110 115 120 aaa ggc ttg aca aat tct tat gtg gga agt aat tat aga cca tca ttt 435 Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn Tyr Arg Pro Ser Phe 125 130 135 ctt cag gat aat gag ctc aga cat tta atc ctt aag act gca gaa ggc 483 Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu Lys Thr Ala Glu Gly 140 145 150 155 ttc tta ttt gtg gtt gga tgt gaa aga gga aaa att ctc ttc gtt tct 531 Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys Ile Leu Phe Val Ser 160 165 170 aag tca gtc tcc aaa ata ctt aat tat gat cag gct agt ttg act gga 579 Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln Ala Ser Leu Thr Gly 175 180 185 caa agc tta ttt gac ttc tta cat cca aaa gat gtt gcc aaa gta aag 627 Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys 190 195 200 gaa caa ctt tct tct ttt gat att tca cca aga gaa aag cta ata gat 675 Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg Glu Lys Leu Ile Asp 205 210 215 gcc aaa act ggt ttg caa gtt cac agt aat ctc cac gct gga agg aca 723 Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu His Ala Gly Arg Thr 220 225 230 235 cgt gtg tat tct ggc tca aga cga tct ttt ttc tgt cgg ata aag agt 771 Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe Cys Arg Ile Lys Ser 240 245 250 tgt aaa atc tct gtc aaa gaa gag cat gga tgc tta ccc aac tca aag 819 Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys Leu Pro Asn Ser Lys 255 260 265 aag aaa gag cac aga aaa ttc tat act atc cat tgc act ggt tac ttg 867 Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His Cys Thr Gly Tyr Leu 270 275 280 aga agc tgg cct cca aat att gtt gga atg gaa gaa gaa agg aac agt 915 Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu Glu Glu Arg Asn Ser 285 290 295 aag aaa gac aac agt aat ttt acc tgc ctt gtg gcc att gga aga tta 963 Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val Ala Ile Gly Arg Leu 300 305 310 315 cag cca tat att gtt cca cag aac agt gga gag att aat gtg aaa cca 1011 Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu Ile Asn Val Lys Pro 320 325 330 act gaa ttt ata acc cgg ttt gca gtg aat gga aaa ttt gtc tat gta 1059 Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly Lys Phe Val Tyr Val 335 340 345 gat caa agg gca aca gcg att tta gga tat ctg cct cag gaa ctt ttg 1107 Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu 350 355 360 gga act tct tgt tat gaa tat ttt cat caa gat gac cac aat aat ttg 1155 Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Asn Asn Leu 365 370 375 act gac aag cac aaa gca gtt cta cag agt aag gag aaa ata ctt aca 1203 Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr 380 385 390 395 gat tcc tac aaa ttc aga gca aaa gat ggc tct ttt gta act tta aaa 1251 Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser Phe Val Thr Leu Lys 400 405 410 agc caa tgg ttt agt ttc aca aat cct tgg aca aaa gaa ctg gaa tat 1299 Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Glu Leu Glu Tyr 415 420 425 att gta tct gtc aac act tta gtt ttg gga cat agt gag cct gga gaa 1347 Ile Val Ser Val Asn Thr Leu Val Leu Gly His Ser Glu Pro Gly Glu 430 435 440 gca tca ttt tta cct tgt agc tct caa tca tca gaa gaa tcc tct aga 1395 Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser Glu Glu Ser Ser Arg 445 450 455 cag tcc tgt atg agt gta cct gga atg tct act gga aca gta ctt ggt 1443 Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr Gly Thr Val Leu Gly 460 465 470 475 gct ggt agt att gga aca gat att gca aat gaa att ctg gat tta cag 1491 Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Ile Leu Asp Leu Gln 480 485 490 agg tta cag tct tct tca tac ctt gat gat tcg agt cca aca ggt tta 1539 Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser Ser Pro Thr Gly Leu 495 500 505 atg aaa gat act cat act gta aac tgc agg agt atg tca aat aag gag 1587 Met Lys Asp Thr His Thr Val Asn Cys Arg Ser Met Ser Asn Lys Glu 510 515 520 ttg ttt cca cca agt cct tct gaa atg ggg gag cta gag gct acc agg 1635 Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu Leu Glu Ala Thr Arg 525 530 535 caa aac cag agt act gtt gct gtc cac agc cat gag cca ctc ctc agt 1683 Gln Asn Gln Ser Thr Val Ala Val His Ser His Glu Pro Leu Leu Ser 540 545 550 555 gat ggt gca cag ttg gat ttc gat gcc cta tgt gac aat gat gac aca 1731 Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys Asp Asn Asp Asp Thr 560 565 570 gcc atg gct gca ttt atg aat tac tta gaa gca gag ggg ggc ctg gga 1779 Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly 575 580 585 gac cct ggg gac ttc agt gac atc cag tgg acc ctc tagc 1819 Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr Leu 590 595 6 599 PRT Homo sapiens 6 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly Glu Val Ala Gly Gly 1 5 10 15 Glu Ala Thr Ala Pro Gly Lys Val Leu Arg Glu Glu Asn Gln Cys Ile 20 25 30 Ala Pro Val Val Ser Ser Arg Val Ser Pro Gly Thr Arg Pro Thr Ala 35 40 45 Met Gly Ser Phe Ser Ser His Met Thr Glu Phe Pro Arg Lys Arg Lys 50 55 60 Gly Ser Asp Ser Asp Pro Ser Gln Glu Ala His Ser Gln Thr Glu Lys 65 70 75 80 Arg Arg Arg Asp Lys Met Asn Asn Leu Ile Glu Glu Leu Ser Ala Met 85 90 95 Ile Pro Gln Cys Asn Pro Met Ala Arg Lys Leu Asp Lys Leu Thr Val 100 105 110 Leu Arg Met Ala Val Gln His Leu Arg Ser Leu Lys Gly Leu Thr Asn 115 120 125 Ser Tyr Val Gly Ser Asn Tyr Arg Pro Ser Phe Leu Gln Asp Asn Glu 130 135 140 Leu Arg His Leu Ile Leu Lys Thr Ala Glu Gly Phe Leu Phe Val Val 145 150 155 160 Gly Cys Glu Arg Gly Lys Ile Leu Phe Val Ser Lys Ser Val Ser Lys 165 170 175 Ile Leu Asn Tyr Asp Gln Ala Ser Leu Thr Gly Gln Ser Leu Phe Asp 180 185 190 Phe Leu His Pro Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser Ser 195 200 205 Phe Asp Ile Ser Pro Arg Glu Lys Leu Ile Asp Ala Lys Thr Gly Leu 210 215 220 Gln Val His Ser Asn Leu His Ala Gly Arg Thr Arg Val Tyr Ser Gly 225 230 235 240 Ser Arg Arg Ser Phe Phe Cys Arg Ile Lys Ser Cys Lys Ile Ser Val 245 250 255 Lys Glu Glu His Gly Cys Leu Pro Asn Ser Lys Lys Lys Glu His Arg 260 265 270 Lys Phe Tyr Thr Ile His Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro 275 280 285 Asn Ile Val Gly Met Glu Glu Glu Arg Asn Ser Lys Lys Asp Asn Ser 290 295 300 Asn Phe Thr Cys Leu Val Ala Ile Gly Arg Leu Gln Pro Tyr Ile Val 305 310 315 320 Pro Gln Asn Ser Gly Glu Ile Asn Val Lys Pro Thr Glu Phe Ile Thr 325 330 335 Arg Phe Ala Val Asn Gly Lys Phe Val Tyr Val Asp Gln Arg Ala Thr 340 345 350 Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr 355 360 365 Glu Tyr Phe His Gln Asp Asp His Asn Asn Leu Thr Asp Lys His Lys 370 375 380 Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe 385 390 395 400 Arg Ala Lys Asp Gly Ser Phe Val Thr Leu Lys Ser Gln Trp Phe Ser 405 410 415 Phe Thr Asn Pro Trp Thr Lys Glu Leu Glu Tyr Ile Val Ser Val Asn 420 425 430 Thr Leu Val Leu Gly His Ser Glu Pro Gly Glu Ala Ser Phe Leu Pro 435 440 445 Cys Ser Ser Gln Ser Ser Glu Glu Ser Ser Arg Gln Ser Cys Met Ser 450 455 460 Val Pro Gly Met Ser Thr Gly Thr Val Leu Gly Ala Gly Ser Ile Gly 465 470 475 480 Thr Asp Ile Ala Asn Glu Ile Leu Asp Leu Gln Arg Leu Gln Ser Ser 485 490 495 Ser Tyr Leu Asp Asp Ser Ser Pro Thr Gly Leu Met Lys Asp Thr His 500 505 510 Thr Val Asn Cys Arg Ser Met Ser Asn Lys Glu Leu Phe Pro Pro Ser 515 520 525 Pro Ser Glu Met Gly Glu Leu Glu Ala Thr Arg Gln Asn Gln Ser Thr 530 535 540 Val Ala Val His Ser His Glu Pro Leu Leu Ser Asp Gly Ala Gln Leu 545 550 555 560 Asp Phe Asp Ala Leu Cys Asp Asn Asp Asp Thr Ala Met Ala Ala Phe 565 570 575 Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe 580 585 590 Ser Asp Ile Gln Trp Thr Leu 595 7 1786 DNA Homo sapiens CDS (19)..(1782) 7 gaccaagtgg ctcctgcg atg gcg gcg gaa gag gag gct gcg gcg gga ggt 51 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly 1 5 10 aaa gtg ttg aga gag gag aac cag tgc att gct cct gtg gtt tcc agc 99 Lys Val Leu Arg Glu Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser 15 20 25 cgc gtg agt cca ggg aca aga cca aca gct atg ggg tct ttc agc tca 147 Arg Val Ser Pro Gly Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser 30 35 40 cac atg aca gag ttt cca cga aaa cgc aaa gga agt gat tca gac cca 195 His Met Thr Glu Phe Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro 45 50 55 tcc caa gaa gct cat agc caa act gaa aag cgg agg aga gat aaa atg 243 Ser Gln Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met 60 65 70 75 aat aac ctg att gaa gaa ctg tct gca atg atc cct cag tgc aac ccc 291 Asn Asn Leu Ile Glu Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro 80 85 90 atg gcg cgt aaa ctg gac aaa ctt aca gtt tta aga atg gct gtt caa 339 Met Ala Arg Lys Leu Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln 95 100 105 cac ttg aga tct tta aaa ggc ttg aca aat tct tat gtg gga agt aat 387 His Leu Arg Ser Leu Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn 110 115 120 tat aga cca tca ttt ctt cag gat aat gag ctc aga cat tta atc ctt 435 Tyr Arg Pro Ser Phe Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu 125 130 135 aag act gca gaa ggc ttc tta ttt gtg gtt gga tgt gaa aga gga aaa 483 Lys Thr Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys 140 145 150 155 att ctc ttc gtt tct aag tca gtc tcc aaa ata ctt aat tat gat cag 531 Ile Leu Phe Val Ser Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln 160 165 170 gct agt ttg act gga caa agc tta ttt gac ttc tta cat cca aaa gat 579 Ala Ser Leu Thr Gly Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp 175 180 185 gtt gcc aaa gta aag gaa caa ctt tct tct ttt gat att tca cca aga 627 Val Ala Lys Val Lys Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg 190 195 200 gaa aag cta ata gat gcc aaa act ggt ttg caa gtt cac agt aat ctc 675 Glu Lys Leu Ile Asp Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu 205 210 215 cac gct gga agg aca cgt gtg tat tct ggc tca aga cga tct ttt ttc 723 His Ala Gly Arg Thr Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe 220 225 230 235 tgt cgg ata aag agt tgt aaa atc tct gtc aaa gaa gag cat gga tgc 771 Cys Arg Ile Lys Ser Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys 240 245 250 tta ccc aac tca aag aag aaa gag cac aga aaa ttc tat act atc cat 819 Leu Pro Asn Ser Lys Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His 255 260 265 tgc act ggt tac ttg aga agc tgg cct cca aat att gtt gga atg gaa 867 Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu 270 275 280 gaa gaa agg aac agt aag aaa gac aac agt aat ttt acc tgc ctt gtg 915 Glu Glu Arg Asn Ser Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val 285 290 295 gcc att gga aga tta cag cca tat att gtt cca cag aac agt gga gag 963 Ala Ile Gly Arg Leu Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu 300 305 310 315 att aat gtg aaa cca act gaa ttt ata acc cgg ttt gca gtg aat gga 1011 Ile Asn Val Lys Pro Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly 320 325 330 aaa ttt gtc tat gta gat caa agg gca aca gcg att tta gga tat ctg 1059 Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu 335 340 345 cct cag gaa ctt ttg gga act tct tgt tat gaa tat ttt cat caa gat 1107 Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp 350 355 360 gac cac aat aat ttg act gac aag cac aaa gca gtt cta cag agt aag 1155 Asp His Asn Asn Leu Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys 365 370 375 gag aaa ata ctt aca gat tcc tac aaa ttc aga gca aaa gat ggc tct 1203 Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser 380 385 390 395 ttt gta act tta aaa agc caa tgg ttt agt ttc aca aat cct tgg aca 1251 Phe Val Thr Leu Lys Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr 400 405 410 aaa gaa ctg gaa tat att gta tct gtc aac act tta gtt ttg gga cat 1299 Lys Glu Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly His 415 420 425 agt gag cct gga gaa gca tca ttt tta cct tgt agc tct caa tca tca 1347 Ser Glu Pro Gly Glu Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser 430 435 440 gaa gaa tcc tct aga cag tcc tgt atg agt gta cct gga atg tct act 1395 Glu Glu Ser Ser Arg Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr 445 450 455 gga aca gta ctt ggt gct ggt agt att gga aca gat att gca aat gaa 1443 Gly Thr Val Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu 460 465 470 475 att ctg gat tta cag agg tta cag tct tct tca tac ctt gat gat tcg 1491 Ile Leu Asp Leu Gln Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser 480 485 490 agt cca aca ggt tta atg aaa gat act cat act gta aac tgc agg agt 1539 Ser Pro Thr Gly Leu Met Lys Asp Thr His Thr Val Asn Cys Arg Ser 495 500 505 atg tca aat aag gag ttg ttt cca cca agt cct tct gaa atg ggg gag 1587 Met Ser Asn Lys Glu Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu 510 515 520 cta gag gct acc agg caa aac cag agt act gtt gct gtc cac agc cat 1635 Leu Glu Ala Thr Arg Gln Asn Gln Ser Thr Val Ala Val His Ser His 525 530 535 gag cca ctc ctc agt gat ggt gca cag ttg gat ttc gat gcc cta tgt 1683 Glu Pro Leu Leu Ser Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys 540 545 550 555 gac aat gat gac aca gcc atg gct gca ttt atg aat tac tta gaa gca 1731 Asp Asn Asp Asp Thr Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala 560 565 570 gag ggg ggc ctg gga gac cct ggg gac ttc agt gac atc cag tgg acc 1779 Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr 575 580 585 ctc tagc 1786 Leu 8 588 PRT Homo sapiens 8 Met Ala Ala Glu Glu Glu Ala Ala Ala Gly Gly Lys Val Leu Arg Glu 1 5 10 15 Glu Asn Gln Cys Ile Ala Pro Val Val Ser Ser Arg Val Ser Pro Gly 20 25 30 Thr Arg Pro Thr Ala Met Gly Ser Phe Ser Ser His Met Thr Glu Phe 35 40 45 Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Pro Ser Gln Glu Ala His 50 55 60 Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn Asn Leu Ile Glu 65 70 75 80 Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro Met Ala Arg Lys Leu 85 90 95 Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln His Leu Arg Ser Leu 100 105 110 Lys Gly Leu Thr Asn Ser Tyr Val Gly Ser Asn Tyr Arg Pro Ser Phe 115 120 125 Leu Gln Asp Asn Glu Leu Arg His Leu Ile Leu Lys Thr Ala Glu Gly 130 135 140 Phe Leu Phe Val Val Gly Cys Glu Arg Gly Lys Ile Leu Phe Val Ser 145 150 155 160 Lys Ser Val Ser Lys Ile Leu Asn Tyr Asp Gln Ala Ser Leu Thr Gly 165 170 175 Gln Ser Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys 180 185 190 Glu Gln Leu Ser Ser Phe Asp Ile Ser Pro Arg Glu Lys Leu Ile Asp 195 200 205 Ala Lys Thr Gly Leu Gln Val His Ser Asn Leu His Ala Gly Arg Thr 210 215 220 Arg Val Tyr Ser Gly Ser Arg Arg Ser Phe Phe Cys Arg Ile Lys Ser 225 230 235 240 Cys Lys Ile Ser Val Lys Glu Glu His Gly Cys Leu Pro Asn Ser Lys 245 250 255 Lys Lys Glu His Arg Lys Phe Tyr Thr Ile His Cys Thr Gly Tyr Leu 260 265 270 Arg Ser Trp Pro Pro Asn Ile Val Gly Met Glu Glu Glu Arg Asn Ser 275 280 285 Lys Lys Asp Asn Ser Asn Phe Thr Cys Leu Val Ala Ile Gly Arg Leu 290 295 300 Gln Pro Tyr Ile Val Pro Gln Asn Ser Gly Glu Ile Asn Val Lys Pro 305 310 315 320 Thr Glu Phe Ile Thr Arg Phe Ala Val Asn Gly Lys Phe Val Tyr Val 325 330 335 Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu 340 345 350 Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Asn Asn Leu 355 360 365 Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr 370 375 380 Asp Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser Phe Val Thr Leu Lys 385 390 395 400 Ser Gln Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Glu Leu Glu Tyr 405 410 415 Ile Val Ser Val Asn Thr Leu Val Leu Gly His Ser Glu Pro Gly Glu 420 425 430 Ala Ser Phe Leu Pro Cys Ser Ser Gln Ser Ser Glu Glu Ser Ser Arg 435 440 445 Gln Ser Cys Met Ser Val Pro Gly Met Ser Thr Gly Thr Val Leu Gly 450 455 460 Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Ile Leu Asp Leu Gln 465 470 475 480 Arg Leu Gln Ser Ser Ser Tyr Leu Asp Asp Ser Ser Pro Thr Gly Leu 485 490 495 Met Lys Asp Thr His Thr Val Asn Cys Arg Ser Met Ser Asn Lys Glu 500 505 510 Leu Phe Pro Pro Ser Pro Ser Glu Met Gly Glu Leu Glu Ala Thr Arg 515 520 525 Gln Asn Gln Ser Thr Val Ala Val His Ser His Glu Pro Leu Leu Ser 530 535 540 Asp Gly Ala Gln Leu Asp Phe Asp Ala Leu Cys Asp Asn Asp Asp Thr 545 550 555 560 Ala Met Ala Ala Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly 565 570 575 Asp Pro Gly Asp Phe Ser Asp Ile Gln Trp Thr Leu 580 585 9 1970 DNA Gallus gallus CDS (102)..(1970) 9 cccccgggcc ggcaggacgg gccgttcctt ctcaccttag ttcacctccc gcatgccgcc 60 ggggccgggg gcgctgtgga gcggggctcg ggccgcccgc c atg gcc gag gca gga 116 Met Ala Glu Ala Gly 1 5 gtg ggg agc gcc gag ggg gca gag gag gag cgg cgg gcc gtt gaa gag 164 Val Gly Ser Ala Glu Gly Ala Glu Glu Glu Arg Arg Ala Val Glu Glu 10 15 20 aat ttt cca gta gat gga aac tcg tgc att gct tct gga gtc ccc agc 212 Asn Phe Pro Val Asp Gly Asn Ser Cys Ile Ala Ser Gly Val Pro Ser 25 30 35 ctc atg aat cca ata act aag cct gct acc act tct ttc aac aat tct 260 Leu Met Asn Pro Ile Thr Lys Pro Ala Thr Thr Ser Phe Asn Asn Ser 40 45 50 gtg gtt gag att cca agg aag cgc aaa gga agt gat tct gat aac cag 308 Val Val Glu Ile Pro Arg Lys Arg Lys Gly Ser Asp Ser Asp Asn Gln 55 60 65 gat aca gtt gaa gtt gat ggg gat cct cag aaa agg aat gaa gat gaa 356 Asp Thr Val Glu Val Asp Gly Asp Pro Gln Lys Arg Asn Glu Asp Glu 70 75 80 85 gaa cat ctt aag ata aaa gat ttc aga gag gcc cac agt caa aca gag 404 Glu His Leu Lys Ile Lys Asp Phe Arg Glu Ala His Ser Gln Thr Glu 90 95 100 aaa cga aga aga gac aaa atg aat aat ttg ata gag gaa ttg tct gct 452 Lys Arg Arg Arg Asp Lys Met Asn Asn Leu Ile Glu Glu Leu Ser Ala 105 110 115 atg ata cct cag tgc aat cct atg gca cga aag cta gac aag ctt aca 500 Met Ile Pro Gln Cys Asn Pro Met Ala Arg Lys Leu Asp Lys Leu Thr 120 125 130 gta tta cgg atg gca gtg caa cac tta aaa tct ttg aaa ggt tcc act 548 Val Leu Arg Met Ala Val Gln His Leu Lys Ser Leu Lys Gly Ser Thr 135 140 145 agc tct tac acc gaa gtc cgg tat aaa cct tcg ttt tta aag gat gat 596 Ser Ser Tyr Thr Glu Val Arg Tyr Lys Pro Ser Phe Leu Lys Asp Asp 150 155 160 165 gag ctc aga cag tta atc ctt agg gct gcg gat gga ttc cta ttt gtg 644 Glu Leu Arg Gln Leu Ile Leu Arg Ala Ala Asp Gly Phe Leu Phe Val 170 175 180 gtt gga tgt aac aga gga aaa att ctg ttt gtc tca gaa tca gtt tgc 692 Val Gly Cys Asn Arg Gly Lys Ile Leu Phe Val Ser Glu Ser Val Cys 185 190 195 aaa ata ctt aat tat gat cag acc agt tta att gga caa agt ttg ttt 740 Lys Ile Leu Asn Tyr Asp Gln Thr Ser Leu Ile Gly Gln Ser Leu Phe 200 205 210 gat tac ttg cat cca aaa gat gtt gcc aaa gtt aag gag caa ctt tca 788 Asp Tyr Leu His Pro Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser 215 220 225 tct tca gat gtc tct ccc aga gaa aag ctt gta gat ggc aaa act ggc 836 Ser Ser Asp Val Ser Pro Arg Glu Lys Leu Val Asp Gly Lys Thr Gly 230 235 240 245 ttg caa gta cat aca gat ttt caa gct gga cca gct cga ctg aat tct 884 Leu Gln Val His Thr Asp Phe Gln Ala Gly Pro Ala Arg Leu Asn Ser 250 255 260 ggt gct cga cgt tcc ttc ttc tgt cgg ata aaa tgt agt agg acc aca 932 Gly Ala Arg Arg Ser Phe Phe Cys Arg Ile Lys Cys Ser Arg Thr Thr 265 270 275 gtc aaa gaa gag aag gag tgc tta ccc aac cca aag aag aaa gat cac 980 Val Lys Glu Glu Lys Glu Cys Leu Pro Asn Pro Lys Lys Lys Asp His 280 285 290 aga aag tat tgt acc att cac tgt act gga tat atg aag aac tgg cct 1028 Arg Lys Tyr Cys Thr Ile His Cys Thr Gly Tyr Met Lys Asn Trp Pro 295 300 305 cct agc gag gtg gga gtg gaa gag gaa aac gat gta gaa aag aac agt 1076 Pro Ser Glu Val Gly Val Glu Glu Glu Asn Asp Val Glu Lys Asn Ser 310 315 320 325 agt aac ttt aac tgt ctc gtt gca att ggg agg tta cac cct tac att 1124 Ser Asn Phe Asn Cys Leu Val Ala Ile Gly Arg Leu His Pro Tyr Ile 330 335 340 gtt cca caa aag agt gga gag ata aaa gtc aaa gca aca gaa ttt gtt 1172 Val Pro Gln Lys Ser Gly Glu Ile Lys Val Lys Ala Thr Glu Phe Val 345 350 355 aca cga ttt gcc atg gat gga aaa ttt gtt tat gta gat cag cgt gca 1220 Thr Arg Phe Ala Met Asp Gly Lys Phe Val Tyr Val Asp Gln Arg Ala 360 365 370 aca gca att tta ggg tat ctg cca caa gag ctt cta gga act tct tgt 1268 Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys 375 380 385 tac gag tac tgc cat caa gat gat cac aat cat cta gct gaa aaa cat 1316 Tyr Glu Tyr Cys His Gln Asp Asp His Asn His Leu Ala Glu Lys His 390 395 400 405 aaa gaa gtg ctg cag aat aaa gaa aaa gta ttt aca aat tcc tac aaa 1364 Lys Glu Val Leu Gln Asn Lys Glu Lys Val Phe Thr Asn Ser Tyr Lys 410 415 420 ttt aga gca aaa gat gga agt ttt att act tta aag agt caa tgg ttt 1412 Phe Arg Ala Lys Asp Gly Ser Phe Ile Thr Leu Lys Ser Gln Trp Phe 425 430 435 agt ttc atg aat ccc tgg acc aag gaa ctg gag tac att gta tca aac 1460 Ser Phe Met Asn Pro Trp Thr Lys Glu Leu Glu Tyr Ile Val Ser Asn 440 445 450 aac act gta gta tta ggt cac aat gag tct gct gaa gaa cag gtc tcc 1508 Asn Thr Val Val Leu Gly His Asn Glu Ser Ala Glu Glu Gln Val Ser 455 460 465 tat ggt tcc cag cct gca gaa ggt gct gta aaa cag tct tta gtg agt 1556 Tyr Gly Ser Gln Pro Ala Glu Gly Ala Val Lys Gln Ser Leu Val Ser 470 475 480 485 gta cct gga atg tcc tct gga aca gtt ctt ggt gct gga agt ata gga 1604 Val Pro Gly Met Ser Ser Gly Thr Val Leu Gly Ala Gly Ser Ile Gly 490 495 500 act gaa att gca aat gaa ata tta gaa tta caa agg ttg cat tct tca 1652 Thr Glu Ile Ala Asn Glu Ile Leu Glu Leu Gln Arg Leu His Ser Ser 505 510 515 ccg cct ggg gag tta agt cca tca cat ctc ttg aga aag tca cca tct 1700 Pro Pro Gly Glu Leu Ser Pro Ser His Leu Leu Arg Lys Ser Pro Ser 520 525 530 cca gct tta act gta aac tgc agc aat gtg ccg aat aaa gag ttg att 1748 Pro Ala Leu Thr Val Asn Cys Ser Asn Val Pro Asn Lys Glu Leu Ile 535 540 545 cag tta tgt cct tca gaa gca gaa gtt ctg gag act tca gaa caa aac 1796 Gln Leu Cys Pro Ser Glu Ala Glu Val Leu Glu Thr Ser Glu Gln Asn 550 555 560 565 caa ggt gct att cca ttc ccc agt aat gag cct ctc ctc ggt ggt aat 1844 Gln Gly Ala Ile Pro Phe Pro Ser Asn Glu Pro Leu Leu Gly Gly Asn 570 575 580 tct cag ctg gac ttt gca ata tgt gaa aat gat gac act gcc atg act 1892 Ser Gln Leu Asp Phe Ala Ile Cys Glu Asn Asp Asp Thr Ala Met Thr 585 590 595 gct ctt atg aat tac ttg gag gcc gat gga gga ctt ggg gat cca gct 1940 Ala Leu Met Asn Tyr Leu Glu Ala Asp Gly Gly Leu Gly Asp Pro Ala 600 605 610 gaa ctc agt gac atc caa tgg gct ctc tag 1970 Glu Leu Ser Asp Ile Gln Trp Ala Leu 615 620 10 622 PRT Gallus gallus 10 Met Ala Glu Ala Gly Val Gly Ser Ala Glu Gly Ala Glu Glu Glu Arg 1 5 10 15 Arg Ala Val Glu Glu Asn Phe Pro Val Asp Gly Asn Ser Cys Ile Ala 20 25 30 Ser Gly Val Pro Ser Leu Met Asn Pro Ile Thr Lys Pro Ala Thr Thr 35 40 45 Ser Phe Asn Asn Ser Val Val Glu Ile Pro Arg Lys Arg Lys Gly Ser 50 55 60 Asp Ser Asp Asn Gln Asp Thr Val Glu Val Asp Gly Asp Pro Gln Lys 65 70 75 80 Arg Asn Glu Asp Glu Glu His Leu Lys Ile Lys Asp Phe Arg Glu Ala 85 90 95 His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn Asn Leu Ile 100 105 110 Glu Glu Leu Ser Ala Met Ile Pro Gln Cys Asn Pro Met Ala Arg Lys 115 120 125 Leu Asp Lys Leu Thr Val Leu Arg Met Ala Val Gln His Leu Lys Ser 130 135 140 Leu Lys Gly Ser Thr Ser Ser Tyr Thr Glu Val Arg Tyr Lys Pro Ser 145 150 155 160 Phe Leu Lys Asp Asp Glu Leu Arg Gln Leu Ile Leu Arg Ala Ala Asp 165 170 175 Gly Phe Leu Phe Val Val Gly Cys Asn Arg Gly Lys Ile Leu Phe Val 180 185 190 Ser Glu Ser Val Cys Lys Ile Leu Asn Tyr Asp Gln Thr Ser Leu Ile 195 200 205 Gly Gln Ser Leu Phe Asp Tyr Leu His Pro Lys Asp Val Ala Lys Val 210 215 220 Lys Glu Gln Leu Ser Ser Ser Asp Val Ser Pro Arg Glu Lys Leu Val 225 230 235 240 Asp Gly Lys Thr Gly Leu Gln Val His Thr Asp Phe Gln Ala Gly Pro 245 250 255 Ala Arg Leu Asn Ser Gly Ala Arg Arg Ser Phe Phe Cys Arg Ile Lys 260 265 270 Cys Ser Arg Thr Thr Val Lys Glu Glu Lys Glu Cys Leu Pro Asn Pro 275 280 285 Lys Lys Lys Asp His Arg Lys Tyr Cys Thr Ile His Cys Thr Gly Tyr 290 295 300 Met Lys Asn Trp Pro Pro Ser Glu Val Gly Val Glu Glu Glu Asn Asp 305 310 315 320 Val Glu Lys Asn Ser Ser Asn Phe Asn Cys Leu Val Ala Ile Gly Arg 325 330 335 Leu His Pro Tyr Ile Val Pro Gln Lys Ser Gly Glu Ile Lys Val Lys 340 345 350 Ala Thr Glu Phe Val Thr Arg Phe Ala Met Asp Gly Lys Phe Val Tyr 355 360 365 Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu 370 375 380 Leu Gly Thr Ser Cys Tyr Glu Tyr Cys His Gln Asp Asp His Asn His 385 390 395 400 Leu Ala Glu Lys His Lys Glu Val Leu Gln Asn Lys Glu Lys Val Phe 405 410 415 Thr Asn Ser Tyr Lys Phe Arg Ala Lys Asp Gly Ser Phe Ile Thr Leu 420 425 430 Lys Ser Gln Trp Phe Ser Phe Met Asn Pro Trp Thr Lys Glu Leu Glu 435 440 445 Tyr Ile Val Ser Asn Asn Thr Val Val Leu Gly His Asn Glu Ser Ala 450 455 460 Glu Glu Gln Val Ser Tyr Gly Ser Gln Pro Ala Glu Gly Ala Val Lys 465 470 475 480 Gln Ser Leu Val Ser Val Pro Gly Met Ser Ser Gly Thr Val Leu Gly 485 490 495 Ala Gly Ser Ile Gly Thr Glu Ile Ala Asn Glu Ile Leu Glu Leu Gln 500 505 510 Arg Leu His Ser Ser Pro Pro Gly Glu Leu Ser Pro Ser His Leu Leu 515 520 525 Arg Lys Ser Pro Ser Pro Ala Leu Thr Val Asn Cys Ser Asn Val Pro 530 535 540 Asn Lys Glu Leu Ile Gln Leu Cys Pro Ser Glu Ala Glu Val Leu Glu 545 550 555 560 Thr Ser Glu Gln Asn Gln Gly Ala Ile Pro Phe Pro Ser Asn Glu Pro 565 570 575 Leu Leu Gly Gly Asn Ser Gln Leu Asp Phe Ala Ile Cys Glu Asn Asp 580 585 590 Asp Thr Ala Met Thr Ala Leu Met Asn Tyr Leu Glu Ala Asp Gly Gly 595 600 605 Leu Gly Asp Pro Ala Glu Leu Ser Asp Ile Gln Trp Ala Leu 610 615 620 11 1752 DNA Mus musculus CDS (12)..(1748) 11 ggtcgaccac c atg gag ttt cca agg aaa cgc aga ggc aga gat tcc cag 50 Met Glu Phe Pro Arg Lys Arg Arg Gly Arg Asp Ser Gln 1 5 10 cca ctc cag tca gaa ttc atg aca gac aca aca gtg gaa agt ctt ccc 98 Pro Leu Gln Ser Glu Phe Met Thr Asp Thr Thr Val Glu Ser Leu Pro 15 20 25 cag aat ccc ttt gcc tct ctt ctt tca aca aga aca gga gta tca gcg 146 Gln Asn Pro Phe Ala Ser Leu Leu Ser Thr Arg Thr Gly Val Ser Ala 30 35 40 45 ccc agt ggc atc agg gaa gct cac agc cag atg gaa aag cgt cgg aga 194 Pro Ser Gly Ile Arg Glu Ala His Ser Gln Met Glu Lys Arg Arg Arg 50 55 60 gac aag atg aac cat ctg att cag aaa ctg tca tct atg atc cct cca 242 Asp Lys Met Asn His Leu Ile Gln Lys Leu Ser Ser Met Ile Pro Pro 65 70 75 cac atc ccc acg gcc cac aaa ctg gac aag ctc agc gtc ttg agg agg 290 His Ile Pro Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg 80 85 90 gcg gtg cag tac ttg agg tct ctg aga ggc atg aca gag ctt tac tta 338 Ala Val Gln Tyr Leu Arg Ser Leu Arg Gly Met Thr Glu Leu Tyr Leu 95 100 105 gga gaa aac tct aaa cct tca ttt att cag gat aag gaa ctc agt cac 386 Gly Glu Asn Ser Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His 110 115 120 125 tta atc ctc aag gca gca gaa ggc ttc ctg ttt gtg gtt gga tgc gaa 434 Leu Ile Leu Lys Ala Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu 130 135 140 aga ggg aga att ttt tac gtt tct aag tct gtc tcc aaa aca ctg cgt 482 Arg Gly Arg Ile Phe Tyr Val Ser Lys Ser Val Ser Lys Thr Leu Arg 145 150 155 tat gat cag gct agc ttg ata gga cag aat ttg ttt gac ttc tta cac 530 Tyr Asp Gln Ala Ser Leu Ile Gly Gln Asn Leu Phe Asp Phe Leu His 160 165 170 cca aaa gac gtc gcc aaa gta aag gaa caa ctt tct tgt gat ggt tca 578 Pro Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Gly Ser 175 180 185 cca aga gag aaa cct ata gac acc aaa acc tct cag gtt tac agt cac 626 Pro Arg Glu Lys Pro Ile Asp Thr Lys Thr Ser Gln Val Tyr Ser His 190 195 200 205 ccc tac act ggg cga cca cgc atg cat tct ggc tcc aga cga tct ttc 674 Pro Tyr Thr Gly Arg Pro Arg Met His Ser Gly Ser Arg Arg Ser Phe 210 215 220 ttc ttt aga atg aag agc tgt acc gtc cct gtc aaa gaa gag cag cca 722 Phe Phe Arg Met Lys Ser Cys Thr Val Pro Val Lys Glu Glu Gln Pro 225 230 235 tgc tcg tcc tgc tca aag aag aaa gac cat aga aaa ttc cac acc gtc 770 Cys Ser Ser Cys Ser Lys Lys Lys Asp His Arg Lys Phe His Thr Val 240 245 250 cat tgc act gga tac ttg aga agc tgg cct ctg aat gtt gtt ggc atg 818 His Cys Thr Gly Tyr Leu Arg Ser Trp Pro Leu Asn Val Val Gly Met 255 260 265 gag aaa gag tcg ggt ggt ggg aag gac agc ggt cct ctt acc tgc ctt 866 Glu Lys Glu Ser Gly Gly Gly Lys Asp Ser Gly Pro Leu Thr Cys Leu 270 275 280 285 gtg gct atg gga cgg ttg cat cca tac att gtc cct caa aag agt ggc 914 Val Ala Met Gly Arg Leu His Pro Tyr Ile Val Pro Gln Lys Ser Gly 290 295 300 aag atc aac gtg aga ccg gct gag ttc ata act cgc ttc gca atg aac 962 Lys Ile Asn Val Arg Pro Ala Glu Phe Ile Thr Arg Phe Ala Met Asn 305 310 315 ggg aaa ttc gtc tat gtt gac caa agg gca acg gca att tta gga tac 1010 Gly Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr 320 325 330 ctg cct cag gaa ctt ttg gga act tca tgt tat gaa tat ttt cat cag 1058 Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln 335 340 345 gat gac cac agt agt ttg act gac aag cac aaa gca gtt ctg cag agt 1106 Asp Asp His Ser Ser Leu Thr Asp Lys His Lys Ala Val Leu Gln Ser 350 355 360 365 aag gag aaa ata ctt aca gac tca tac aaa ttc aga gtg aag gat ggt 1154 Lys Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Val Lys Asp Gly 370 375 380 gcc ttc gtg act ctg aag agt gag tgg ttc agc ttc aca aac cct tgg 1202 Ala Phe Val Thr Leu Lys Ser Glu Trp Phe Ser Phe Thr Asn Pro Trp 385 390 395 acc aaa gag ctg gag tac att gtg tct gtc aac aca ttg gtt ttg ggg 1250 Thr Lys Glu Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly 400 405 410 cgc agt gag acc agg ctg tct ttg ctt cat tgc ggc ggc agc agc cag 1298 Arg Ser Glu Thr Arg Leu Ser Leu Leu His Cys Gly Gly Ser Ser Gln 415 420 425 tcc tcc gaa gac tca ttt aga caa tcc tgc atc aat gtg ccc ggt gta 1346 Ser Ser Glu Asp Ser Phe Arg Gln Ser Cys Ile Asn Val Pro Gly Val 430 435 440 445 tcc acg ggg acc gtc ctt ggt gct ggg agt att gga aca gat att gca 1394 Ser Thr Gly Thr Val Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala 450 455 460 aat gag gtt ctg agt tta cag aga tta cac tct tca tcc cca gaa gat 1442 Asn Glu Val Leu Ser Leu Gln Arg Leu His Ser Ser Ser Pro Glu Asp 465 470 475 gca agc cct tca gaa gaa gtg aga gat gac tgc agt gta aat ggt ggg 1490 Ala Ser Pro Ser Glu Glu Val Arg Asp Asp Cys Ser Val Asn Gly Gly 480 485 490 aat gcc tat ggg cct gca tcc act agg gag cct ttt gca gtg agc cct 1538 Asn Ala Tyr Gly Pro Ala Ser Thr Arg Glu Pro Phe Ala Val Ser Pro 495 500 505 tct gaa aca gag gtc ctg gag gct gcc agg caa cac cag agc act gaa 1586 Ser Glu Thr Glu Val Leu Glu Ala Ala Arg Gln His Gln Ser Thr Glu 510 515 520 525 ccc gcc cac cct cac gga cca ctt ccc ggt gac agt gcc cag ctg ggt 1634 Pro Ala His Pro His Gly Pro Leu Pro Gly Asp Ser Ala Gln Leu Gly 530 535 540 ttt gat gtc ctg tgt gac agt gac agc ata gac atg gct gca ttc atg 1682 Phe Asp Val Leu Cys Asp Ser Asp Ser Ile Asp Met Ala Ala Phe Met 545 550 555 aat tac ctc gaa gca gag ggg ggc ctg ggt gac cct ggg gac ttc agt 1730 Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe Ser 560 565 570 gac atc cag tgg gca ctc tagc 1752 Asp Ile Gln Trp Ala Leu 575 12 579 PRT Mus musculus 12 Met Glu Phe Pro Arg Lys Arg Arg Gly Arg Asp Ser Gln Pro Leu Gln 1 5 10 15 Ser Glu Phe Met Thr Asp Thr Thr Val Glu Ser Leu Pro Gln Asn Pro 20 25 30 Phe Ala Ser Leu Leu Ser Thr Arg Thr Gly Val Ser Ala Pro Ser Gly 35 40 45 Ile Arg Glu Ala His Ser Gln Met Glu Lys Arg Arg Arg Asp Lys Met 50 55 60 Asn His Leu Ile Gln Lys Leu Ser Ser Met Ile Pro Pro His Ile Pro 65 70 75 80 Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg Ala Val Gln 85 90 95 Tyr Leu Arg Ser Leu Arg Gly Met Thr Glu Leu Tyr Leu Gly Glu Asn 100 105 110 Ser Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His Leu Ile Leu 115 120 125 Lys Ala Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg Gly Arg 130 135 140 Ile Phe Tyr Val Ser Lys Ser Val Ser Lys Thr Leu Arg Tyr Asp Gln 145 150 155 160 Ala Ser Leu Ile Gly Gln Asn Leu Phe Asp Phe Leu His Pro Lys Asp 165 170 175 Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Gly Ser Pro Arg Glu 180 185 190 Lys Pro Ile Asp Thr Lys Thr Ser Gln Val Tyr Ser His Pro Tyr Thr 195 200 205 Gly Arg Pro Arg Met His Ser Gly Ser Arg Arg Ser Phe Phe Phe Arg 210 215 220 Met Lys Ser Cys Thr Val Pro Val Lys Glu Glu Gln Pro Cys Ser Ser 225 230 235 240 Cys Ser Lys Lys Lys Asp His Arg Lys Phe His Thr Val His Cys Thr 245 250 255 Gly Tyr Leu Arg Ser Trp Pro Leu Asn Val Val Gly Met Glu Lys Glu 260 265 270 Ser Gly Gly Gly Lys Asp Ser Gly Pro Leu Thr Cys Leu Val Ala Met 275 280 285 Gly Arg Leu His Pro Tyr Ile Val Pro Gln Lys Ser Gly Lys Ile Asn 290 295 300 Val Arg Pro Ala Glu Phe Ile Thr Arg Phe Ala Met Asn Gly Lys Phe 305 310 315 320 Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln 325 330 335 Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His 340 345 350 Ser Ser Leu Thr Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys 355 360 365 Ile Leu Thr Asp Ser Tyr Lys Phe Arg Val Lys Asp Gly Ala Phe Val 370 375 380 Thr Leu Lys Ser Glu Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Glu 385 390 395 400 Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly Arg Ser Glu 405 410 415 Thr Arg Leu Ser Leu Leu His Cys Gly Gly Ser Ser Gln Ser Ser Glu 420 425 430 Asp Ser Phe Arg Gln Ser Cys Ile Asn Val Pro Gly Val Ser Thr Gly 435 440 445 Thr Val Leu Gly Ala Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Val 450 455 460 Leu Ser Leu Gln Arg Leu His Ser Ser Ser Pro Glu Asp Ala Ser Pro 465 470 475 480 Ser Glu Glu Val Arg Asp Asp Cys Ser Val Asn Gly Gly Asn Ala Tyr 485 490 495 Gly Pro Ala Ser Thr Arg Glu Pro Phe Ala Val Ser Pro Ser Glu Thr 500 505 510 Glu Val Leu Glu Ala Ala Arg Gln His Gln Ser Thr Glu Pro Ala His 515 520 525 Pro His Gly Pro Leu Pro Gly Asp Ser Ala Gln Leu Gly Phe Asp Val 530 535 540 Leu Cys Asp Ser Asp Ser Ile Asp Met Ala Ala Phe Met Asn Tyr Leu 545 550 555 560 Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe Ser Asp Ile Gln 565 570 575 Trp Ala Leu 13 1676 DNA Mus musculus CDS (12)..(608) 13 ggtcgaccac c atg gag ttt cca agg aaa cgc aga ggc aga gat tcc cag 50 Met Glu Phe Pro Arg Lys Arg Arg Gly Arg Asp Ser Gln 1 5 10 cca ctc cag tca gaa ttc atg aca gac aca aca gtg gaa agt ctt ccc 98 Pro Leu Gln Ser Glu Phe Met Thr Asp Thr Thr Val Glu Ser Leu Pro 15 20 25 cag aat ccc ttt gcc tct ctt ctt tca aca aga aca gga gta tca gcg 146 Gln Asn Pro Phe Ala Ser Leu Leu Ser Thr Arg Thr Gly Val Ser Ala 30 35 40 45 ccc agt ggc atc agg gaa gct cac agc cag atg gaa aag cgt cgg aga 194 Pro Ser Gly Ile Arg Glu Ala His Ser Gln Met Glu Lys Arg Arg Arg 50 55 60 gac aag atg aac cat ctg att cag aaa ctg tca tct atg atc cct cca 242 Asp Lys Met Asn His Leu Ile Gln Lys Leu Ser Ser Met Ile Pro Pro 65 70 75 cac atc ccc acg gcc cac aaa ctg gac aag ctc agc gtc ttg agg agg 290 His Ile Pro Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg 80 85 90 gcg gtg cag tac ttg agg tct ctg aga ggc atg aca gag ctt tac tta 338 Ala Val Gln Tyr Leu Arg Ser Leu Arg Gly Met Thr Glu Leu Tyr Leu 95 100 105 gga gaa aac tct aaa cct tca ttt att cag gat aag gaa ctc agt cac 386 Gly Glu Asn Ser Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His 110 115 120 125 tta atc ctc aag gca gca gaa ggc ttc ctg ttt gtg gtt gga tgc gaa 434 Leu Ile Leu Lys Ala Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu 130 135 140 aga ggg aga att ttt tac gtt tct aag tct gtc tcc aaa aca ctg cgt 482 Arg Gly Arg Ile Phe Tyr Val Ser Lys Ser Val Ser Lys Thr Leu Arg 145 150 155 tat gat cag gct agc ttg ata gga cag aat ttg ttt gac ttc tta cac 530 Tyr Asp Gln Ala Ser Leu Ile Gly Gln Asn Leu Phe Asp Phe Leu His 160 165 170 cca aaa gac gtc gcc aaa gta aag gaa caa ctt tct tgt gat ggt tca 578 Pro Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Gly Ser 175 180 185 cca aga gag aaa cct ata gac acc aaa aaa tgaagagctg taccgtccct 628 Pro Arg Glu Lys Pro Ile Asp Thr Lys Lys 190 195 gtcaaagaag agcagccatg ctcgtcctgc tcaaagaaga aagaccatag aaaattccac 688 accgtccatt gcactggata cttgagaagc tggcctctga atgttgttgg catggagaaa 748 gagtcgggtg gtgggaagga cagcggtcct cttacctgcc ttgtggctat gggacggttg 808 catccataca ttgtccctca aaagagtggc aagatcaacg tgagaccggc tgagttcata 868 actcgcttcg caatgaacgg gaaattcgtc tatgttgacc aaagggcaac ggcaatttta 928 ggatacctgc ctcaggaact tttgggaact tcatgttatg aatattttca tcaggatgac 988 cacagtagtt tgactgacaa gcacaaagca gttctgcaga gtaaggagaa aatacttaca 1048 gactcataca aattcagagt gaaggatggt gccttcgtga ctctgaagag tgagtggttc 1108 agcttcacaa acccttggac caaagagctg gagtacattg tgtctgtcaa cacattggtt 1168 ttggggcgca gtgagaccag gctgtctttg cttcattgcg gcggcagcag ccagtcctcc 1228 gaagactcat ttagacaatc ctgcatcaat gtgcccggtg tatccacggg gaccgtcctt 1288 ggtgctggga gtattggaac agatattgca aatgaggttc tgagtttaca gagattacac 1348 tcttcatccc cagaagatgc aagcccttca gaagaagtga gagatgactg cagtgtaaat 1408 ggtgggaatg cctatgggcc tgcatccact agggagcctt ttgcagtgag cccttctgaa 1468 acagaggtcc tggaggctgc caggcaacac cagagcactg aacccgccca ccctcacgga 1528 ccacttcccg gtgacagtgc ccagctgggt tttgatgtcc tgtgtgacag tgacagcata 1588 gacatggctg cattcatgaa ttacctcgaa gcagaggggg gcctgggtga ccctggggac 1648 ttcagtgaca tccagtgggc actctagc 1676 14 199 PRT Mus musculus 14 Met Glu Phe Pro Arg Lys Arg Arg Gly Arg Asp Ser Gln Pro Leu Gln 1 5 10 15 Ser Glu Phe Met Thr Asp Thr Thr Val Glu Ser Leu Pro Gln Asn Pro 20 25 30 Phe Ala Ser Leu Leu Ser Thr Arg Thr Gly Val Ser Ala Pro Ser Gly 35 40 45 Ile Arg Glu Ala His Ser Gln Met Glu Lys Arg Arg Arg Asp Lys Met 50 55 60 Asn His Leu Ile Gln Lys Leu Ser Ser Met Ile Pro Pro His Ile Pro 65 70 75 80 Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg Ala Val Gln 85 90 95 Tyr Leu Arg Ser Leu Arg Gly Met Thr Glu Leu Tyr Leu Gly Glu Asn 100 105 110 Ser Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His Leu Ile Leu 115 120 125 Lys Ala Ala Glu Gly Phe Leu Phe Val Val Gly Cys Glu Arg Gly Arg 130 135 140 Ile Phe Tyr Val Ser Lys Ser Val Ser Lys Thr Leu Arg Tyr Asp Gln 145 150 155 160 Ala Ser Leu Ile Gly Gln Asn Leu Phe Asp Phe Leu His Pro Lys Asp 165 170 175 Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Gly Ser Pro Arg Glu 180 185 190 Lys Pro Ile Asp Thr Lys Lys 195 15 1748 DNA Rattus norvegicus CDS (33)..(1745) 15 tcacagctcc tggacctgtc agctctcttg ca atg gag ttg cca agg aaa cgt 53 Met Glu Leu Pro Arg Lys Arg 1 5 aga aga agt gat tca gag ctg ctc cag tca gaa ttc agg aca gat gca 101 Arg Arg Ser Asp Ser Glu Leu Leu Gln Ser Glu Phe Arg Thr Asp Ala 10 15 20 atg gtg gaa aac ctt ccc cgg agt ccc ttt acc tct gtt ctt tca aca 149 Met Val Glu Asn Leu Pro Arg Ser Pro Phe Thr Ser Val Leu Ser Thr 25 30 35 aga aca gga gta gca gtg ccc aat ggc atc agg gaa gct cac agc cag 197 Arg Thr Gly Val Ala Val Pro Asn Gly Ile Arg Glu Ala His Ser Gln 40 45 50 55 aca gaa aag cgt cgg aga gac aag atg aac cat ctg att tgg aaa ctg 245 Thr Glu Lys Arg Arg Arg Asp Lys Met Asn His Leu Ile Trp Lys Leu 60 65 70 tca tct atg atc cct cca cac atc ccc aca gcc cac aaa ctg gac aaa 293 Ser Ser Met Ile Pro Pro His Ile Pro Thr Ala His Lys Leu Asp Lys 75 80 85 ctg agc gtc ctg agg agg gca gtg cag tac ttg agg tct cag aga ggc 341 Leu Ser Val Leu Arg Arg Ala Val Gln Tyr Leu Arg Ser Gln Arg Gly 90 95 100 atg aca gag ttt tat tta gga gaa aat gct aaa cct tca ttt att cag 389 Met Thr Glu Phe Tyr Leu Gly Glu Asn Ala Lys Pro Ser Phe Ile Gln 105 110 115 gat aag gaa ctc agc cac tta atc ctc aag gca gca gaa ggc ttc cta 437 Asp Lys Glu Leu Ser His Leu Ile Leu Lys Ala Ala Glu Gly Phe Leu 120 125 130 135 ctt gtg gtt gga tgt gaa gga ggg aga att ctt ttc gtt tct aag tct 485 Leu Val Val Gly Cys Glu Gly Gly Arg Ile Leu Phe Val Ser Lys Ser 140 145 150 gtc tcc aaa acg ctg cat tat gat cag gct agt ttg atg gga cag aac 533 Val Ser Lys Thr Leu His Tyr Asp Gln Ala Ser Leu Met Gly Gln Asn 155 160 165 ttg ttt gac ttc tta cac cca aaa gat gtc gcc aaa gta aag gaa caa 581 Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys Glu Gln 170 175 180 ctt tct tgt gat gtt tca ctg aga gag aaa ccc ata ggc acc aaa acc 629 Leu Ser Cys Asp Val Ser Leu Arg Glu Lys Pro Ile Gly Thr Lys Thr 185 190 195 tct cct cag gtt cac agt cac tcc cat att ggg cga tca cgc gtg cat 677 Ser Pro Gln Val His Ser His Ser His Ile Gly Arg Ser Arg Val His 200 205 210 215 tct ggc tcc aga cga tct ttc ttc ttt aga atg aag agc agc tgt aca 725 Ser Gly Ser Arg Arg Ser Phe Phe Phe Arg Met Lys Ser Ser Cys Thr 220 225 230 gtc ccc gtc aaa gaa gag caa cga tgc tcg tcc tgt tca aag aag aaa 773 Val Pro Val Lys Glu Glu Gln Arg Cys Ser Ser Cys Ser Lys Lys Lys 235 240 245 gac cag aga aaa ttc cac acc atc cat tgc act gga tac ttg aga agc 821 Asp Gln Arg Lys Phe His Thr Ile His Cys Thr Gly Tyr Leu Arg Ser 250 255 260 tgg cca ccg aat gtt gtg ggc acg gag aaa gag atg ggc agt ggg aaa 869 Trp Pro Pro Asn Val Val Gly Thr Glu Lys Glu Met Gly Ser Gly Lys 265 270 275 gac agt ggt cct ctt acc tgc ctt gtg gct atg gga cgg tta cag cca 917 Asp Ser Gly Pro Leu Thr Cys Leu Val Ala Met Gly Arg Leu Gln Pro 280 285 290 295 tat act gtc ccc ccg aag aat ggc aag atc aac gtg aga ccg gct gag 965 Tyr Thr Val Pro Pro Lys Asn Gly Lys Ile Asn Val Arg Pro Ala Glu 300 305 310 ttc ata acc cga ttc gca atg aac ggg aaa ttc gtc tac gtc gac caa 1013 Phe Ile Thr Arg Phe Ala Met Asn Gly Lys Phe Val Tyr Val Asp Gln 315 320 325 agg gca aca gca att tta gga tac ctg cct cag gaa ctt ttg gga act 1061 Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu Gly Thr 330 335 340 tcg tgt tat gaa tat ttt cat cag gat gac cac agt aat ttg agt gac 1109 Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Ser Asn Leu Ser Asp 345 350 355 aag cac aaa gca gtt ctg cag agt aag gag aaa ata ctt aca gat tca 1157 Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr Asp Ser 360 365 370 375 tac aaa ttc aga gtg aag gat ggc tcc ttt gtg act ctg aag agc aag 1205 Tyr Lys Phe Arg Val Lys Asp Gly Ser Phe Val Thr Leu Lys Ser Lys 380 385 390 tgg ttc agc ttc act aac cct tgg acc aaa aag ctg gag tac atc gtg 1253 Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Lys Leu Glu Tyr Ile Val 395 400 405 tct gtc aac acg ctg gtt ttg ggg cgc agt gag acc gca gta tcc gtg 1301 Ser Val Asn Thr Leu Val Leu Gly Arg Ser Glu Thr Ala Val Ser Val 410 415 420 cct cag tgc cgc agc agc cag tcc tct gaa gac tca ttt aga caa ccc 1349 Pro Gln Cys Arg Ser Ser Gln Ser Ser Glu Asp Ser Phe Arg Gln Pro 425 430 435 tgc gtc agt gtg ccg ggc ata tcc aca ggg acc tta ctt ggc act ggg 1397 Cys Val Ser Val Pro Gly Ile Ser Thr Gly Thr Leu Leu Gly Thr Gly 440 445 450 455 agt att gga aca gat att gca aat gag gtt ctg agt tta cag agg tca 1445 Ser Ile Gly Thr Asp Ile Ala Asn Glu Val Leu Ser Leu Gln Arg Ser 460 465 470 cac tct tca tcc cca gaa gac gca aac cct tca gga gta gtg aga gat 1493 His Ser Ser Ser Pro Glu Asp Ala Asn Pro Ser Gly Val Val Arg Asp 475 480 485 aag cac agt gta aac ttc ggg agc gcc cct gtg ccc gtg tcc act ggg 1541 Lys His Ser Val Asn Phe Gly Ser Ala Pro Val Pro Val Ser Thr Gly 490 495 500 gag ctc ttt gca ctg agt cct gaa aca gag ggc ctg gag gct gcc agg 1589 Glu Leu Phe Ala Leu Ser Pro Glu Thr Glu Gly Leu Glu Ala Ala Arg 505 510 515 caa cac cag agt tct gag ccc gcc cac tgt cac aaa cca ctc ctc agt 1637 Gln His Gln Ser Ser Glu Pro Ala His Cys His Lys Pro Leu Leu Ser 520 525 530 535 gac agt acc cag ttg ggt ttt gat gcc ctg tgt gac agc gac gac aca 1685 Asp Ser Thr Gln Leu Gly Phe Asp Ala Leu Cys Asp Ser Asp Asp Thr 540 545 550 gcc atg gct aca ttc atg aat tac ctc gaa gca gag ggt ggc ctg ggt 1733 Ala Met Ala Thr Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly 555 560 565 gac cct ggg gac ttc 1748 Asp Pro Gly Asp 570 16 571 PRT Rattus norvegicus 16 Met Glu Leu Pro Arg Lys Arg Arg Arg Ser Asp Ser Glu Leu Leu Gln 1 5 10 15 Ser Glu Phe Arg Thr Asp Ala Met Val Glu Asn Leu Pro Arg Ser Pro 20 25 30 Phe Thr Ser Val Leu Ser Thr Arg Thr Gly Val Ala Val Pro Asn Gly 35 40 45 Ile Arg Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met 50 55 60 Asn His Leu Ile Trp Lys Leu Ser Ser Met Ile Pro Pro His Ile Pro 65 70 75 80 Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg Ala Val Gln 85 90 95 Tyr Leu Arg Ser Gln Arg Gly Met Thr Glu Phe Tyr Leu Gly Glu Asn 100 105 110 Ala Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His Leu Ile Leu 115 120 125 Lys Ala Ala Glu Gly Phe Leu Leu Val Val Gly Cys Glu Gly Gly Arg 130 135 140 Ile Leu Phe Val Ser Lys Ser Val Ser Lys Thr Leu His Tyr Asp Gln 145 150 155 160 Ala Ser Leu Met Gly Gln Asn Leu Phe Asp Phe Leu His Pro Lys Asp 165 170 175 Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Val Ser Leu Arg Glu 180 185 190 Lys Pro Ile Gly Thr Lys Thr Ser Pro Gln Val His Ser His Ser His 195 200 205 Ile Gly Arg Ser Arg Val His Ser Gly Ser Arg Arg Ser Phe Phe Phe 210 215 220 Arg Met Lys Ser Ser Cys Thr Val Pro Val Lys Glu Glu Gln Arg Cys 225 230 235 240 Ser Ser Cys Ser Lys Lys Lys Asp Gln Arg Lys Phe His Thr Ile His 245 250 255 Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro Asn Val Val Gly Thr Glu 260 265 270 Lys Glu Met Gly Ser Gly Lys Asp Ser Gly Pro Leu Thr Cys Leu Val 275 280 285 Ala Met Gly Arg Leu Gln Pro Tyr Thr Val Pro Pro Lys Asn Gly Lys 290 295 300 Ile Asn Val Arg Pro Ala Glu Phe Ile Thr Arg Phe Ala Met Asn Gly 305 310 315 320 Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu 325 330 335 Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp 340 345 350 Asp His Ser Asn Leu Ser Asp Lys His Lys Ala Val Leu Gln Ser Lys 355 360 365 Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Val Lys Asp Gly Ser 370 375 380 Phe Val Thr Leu Lys Ser Lys Trp Phe Ser Phe Thr Asn Pro Trp Thr 385 390 395 400 Lys Lys Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly Arg 405 410 415 Ser Glu Thr Ala Val Ser Val Pro Gln Cys Arg Ser Ser Gln Ser Ser 420 425 430 Glu Asp Ser Phe Arg Gln Pro Cys Val Ser Val Pro Gly Ile Ser Thr 435 440 445 Gly Thr Leu Leu Gly Thr Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu 450 455 460 Val Leu Ser Leu Gln Arg Ser His Ser Ser Ser Pro Glu Asp Ala Asn 465 470 475 480 Pro Ser Gly Val Val Arg Asp Lys His Ser Val Asn Phe Gly Ser Ala 485 490 495 Pro Val Pro Val Ser Thr Gly Glu Leu Phe Ala Leu Ser Pro Glu Thr 500 505 510 Glu Gly Leu Glu Ala Ala Arg Gln His Gln Ser Ser Glu Pro Ala His 515 520 525 Cys His Lys Pro Leu Leu Ser Asp Ser Thr Gln Leu Gly Phe Asp Ala 530 535 540 Leu Cys Asp Ser Asp Asp Thr Ala Met Ala Thr Phe Met Asn Tyr Leu 545 550 555 560 Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp 565 570 17 1646 DNA Rattus norvegicus CDS (33)..(1646) 17 tcacagctcc tggacctgtc agctctcttg ca atg gag ttg cca agg aaa cgt 53 Met Glu Leu Pro Arg Lys Arg 1 5 aga aga agt gat tca gag ctg ctc cag gaa gct cac agc cag aca gaa 101 Arg Arg Ser Asp Ser Glu Leu Leu Gln Glu Ala His Ser Gln Thr Glu 10 15 20 aag cgt cgg aga gac aag atg aac cat ctg att tgg aaa ctg tca tct 149 Lys Arg Arg Arg Asp Lys Met Asn His Leu Ile Trp Lys Leu Ser Ser 25 30 35 atg atc cct cca cac atc ccc aca gcc cac aaa ctg gac aaa ctg agc 197 Met Ile Pro Pro His Ile Pro Thr Ala His Lys Leu Asp Lys Leu Ser 40 45 50 55 gtc ctg agg agg gca gtg cag tac ttg agg tct cag aga ggc atg aca 245 Val Leu Arg Arg Ala Val Gln Tyr Leu Arg Ser Gln Arg Gly Met Thr 60 65 70 gag ttt tat tta gga gaa aat gct aaa cct tca ttt att cag gat aag 293 Glu Phe Tyr Leu Gly Glu Asn Ala Lys Pro Ser Phe Ile Gln Asp Lys 75 80 85 gaa ctc agc cac tta atc ctc aag gca gca gaa ggc ttc cta ctt gtg 341 Glu Leu Ser His Leu Ile Leu Lys Ala Ala Glu Gly Phe Leu Leu Val 90 95 100 gtt gga tgt gaa gga ggg aga att ctt ttc gtt tct aag tct gtc tcc 389 Val Gly Cys Glu Gly Gly Arg Ile Leu Phe Val Ser Lys Ser Val Ser 105 110 115 aaa acg ctg cat tat gat cag gct agt ttg atg gga cag aac ttg ttt 437 Lys Thr Leu His Tyr Asp Gln Ala Ser Leu Met Gly Gln Asn Leu Phe 120 125 130 135 gac ttc tta cac cca aaa gat gtc gcc aaa gta aag gaa caa ctt tct 485 Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys Glu Gln Leu Ser 140 145 150 tgt gat gtt tca ctg aga gag aaa ccc ata ggc acc aaa acc tct cct 533 Cys Asp Val Ser Leu Arg Glu Lys Pro Ile Gly Thr Lys Thr Ser Pro 155 160 165 cag gtt cac agt cac tcc cat att ggg cga tca cgc gtg cat tct ggc 581 Gln Val His Ser His Ser His Ile Gly Arg Ser Arg Val His Ser Gly 170 175 180 tcc aga cga tct ttc ttc ttt aga atg aag agc agc tgt aca gtc ccc 629 Ser Arg Arg Ser Phe Phe Phe Arg Met Lys Ser Ser Cys Thr Val Pro 185 190 195 gtc aaa gaa gag caa cga tgc tcg tcc tgt tca aag aag aaa gac cag 677 Val Lys Glu Glu Gln Arg Cys Ser Ser Cys Ser Lys Lys Lys Asp Gln 200 205 210 215 aga aaa ttc cac acc atc cat tgc act gga tac ttg aga agc tgg cca 725 Arg Lys Phe His Thr Ile His Cys Thr Gly Tyr Leu Arg Ser Trp Pro 220 225 230 ccg aat gtt gtg ggc acg gag aaa gag atg ggc agt ggg aaa gac agt 773 Pro Asn Val Val Gly Thr Glu Lys Glu Met Gly Ser Gly Lys Asp Ser 235 240 245 ggt cct ctt acc tgc ctt gtg gct atg gga cgg tta cag cca tat act 821 Gly Pro Leu Thr Cys Leu Val Ala Met Gly Arg Leu Gln Pro Tyr Thr 250 255 260 gtc ccc ccg aag aat ggc aag atc aac gtg aga ccg gct gag ttc ata 869 Val Pro Pro Lys Asn Gly Lys Ile Asn Val Arg Pro Ala Glu Phe Ile 265 270 275 acc cga ttc gca atg aac ggg aaa ttc gtc tac gtc gac caa agg gca 917 Thr Arg Phe Ala Met Asn Gly Lys Phe Val Tyr Val Asp Gln Arg Ala 280 285 290 295 aca gca att tta gga tac ctg cct cag gaa ctt ttg gga act tcg tgt 965 Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys 300 305 310 tat gaa tat ttt cat cag gat gac cac agt aat ttg agt gac aag cac 1013 Tyr Glu Tyr Phe His Gln Asp Asp His Ser Asn Leu Ser Asp Lys His 315 320 325 aaa gca gtt ctg cag agt aag gag aaa ata ctt aca gat tca tac aaa 1061 Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr Asp Ser Tyr Lys 330 335 340 ttc aga gtg aag gat ggc tcc ttt gtg act ctg aag agc aag tgg ttc 1109 Phe Arg Val Lys Asp Gly Ser Phe Val Thr Leu Lys Ser Lys Trp Phe 345 350 355 agc ttc act aac cct tgg acc aaa aag ctg gag tac atc gtg tct gtc 1157 Ser Phe Thr Asn Pro Trp Thr Lys Lys Leu Glu Tyr Ile Val Ser Val 360 365 370 375 aac acg ctg gtt ttg ggg cgc agt gag acc gca gta tcc gtg cct cag 1205 Asn Thr Leu Val Leu Gly Arg Ser Glu Thr Ala Val Ser Val Pro Gln 380 385 390 tgc cgc agc agc cag tcc tct gaa gac tca ttt aga caa ccc tgc gtc 1253 Cys Arg Ser Ser Gln Ser Ser Glu Asp Ser Phe Arg Gln Pro Cys Val 395 400 405 agt gtg ccg ggc ata tcc aca ggg acc tta ctt ggc act ggg agt att 1301 Ser Val Pro Gly Ile Ser Thr Gly Thr Leu Leu Gly Thr Gly Ser Ile 410 415 420 gga aca gat att gca aat gag gtt ctg agt tta cag agg tca cac tct 1349 Gly Thr Asp Ile Ala Asn Glu Val Leu Ser Leu Gln Arg Ser His Ser 425 430 435 tca tcc cca gaa gac gca aac cct tca gga gta gtg aga gat aag cac 1397 Ser Ser Pro Glu Asp Ala Asn Pro Ser Gly Val Val Arg Asp Lys His 440 445 450 455 agt gta aac ttc ggg agc gcc cct gtg ccc gtg tcc act ggg gag ctc 1445 Ser Val Asn Phe Gly Ser Ala Pro Val Pro Val Ser Thr Gly Glu Leu 460 465 470 ttt gca ctg agt cct gaa aca gag ggc ctg gag gct gcc agg caa cac 1493 Phe Ala Leu Ser Pro Glu Thr Glu Gly Leu Glu Ala Ala Arg Gln His 475 480 485 cag agt tct gag ccc gcc cac tgt cac aaa cca ctc ctc agt gac agt 1541 Gln Ser Ser Glu Pro Ala His Cys His Lys Pro Leu Leu Ser Asp Ser 490 495 500 acc cag ttg ggt ttt gat gcc ctg tgt gac agc gac gac aca gcc atg 1589 Thr Gln Leu Gly Phe Asp Ala Leu Cys Asp Ser Asp Asp Thr Ala Met 505 510 515 gct aca ttc atg aat tac ctc gaa gca gag ggt ggc ctg ggt gac cct 1637 Ala Thr Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu Gly Asp Pro 520 525 530 535 ggg gac ttc 1646 Gly Asp Phe 18 538 PRT Rattus norvegicus 18 Met Glu Leu Pro Arg Lys Arg Arg Arg Ser Asp Ser Glu Leu Leu Gln 1 5 10 15 Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met Asn His 20 25 30 Leu Ile Trp Lys Leu Ser Ser Met Ile Pro Pro His Ile Pro Thr Ala 35 40 45 His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg Ala Val Gln Tyr Leu 50 55 60 Arg Ser Gln Arg Gly Met Thr Glu Phe Tyr Leu Gly Glu Asn Ala Lys 65 70 75 80 Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His Leu Ile Leu Lys Ala 85 90 95 Ala Glu Gly Phe Leu Leu Val Val Gly Cys Glu Gly Gly Arg Ile Leu 100 105 110 Phe Val Ser Lys Ser Val Ser Lys Thr Leu His Tyr Asp Gln Ala Ser 115 120 125 Leu Met Gly Gln Asn Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala 130 135 140 Lys Val Lys Glu Gln Leu Ser Cys Asp Val Ser Leu Arg Glu Lys Pro 145 150 155 160 Ile Gly Thr Lys Thr Ser Pro Gln Val His Ser His Ser His Ile Gly 165 170 175 Arg Ser Arg Val His Ser Gly Ser Arg Arg Ser Phe Phe Phe Arg Met 180 185 190 Lys Ser Ser Cys Thr Val Pro Val Lys Glu Glu Gln Arg Cys Ser Ser 195 200 205 Cys Ser Lys Lys Lys Asp Gln Arg Lys Phe His Thr Ile His Cys Thr 210 215 220 Gly Tyr Leu Arg Ser Trp Pro Pro Asn Val Val Gly Thr Glu Lys Glu 225 230 235 240 Met Gly Ser Gly Lys Asp Ser Gly Pro Leu Thr Cys Leu Val Ala Met 245 250 255 Gly Arg Leu Gln Pro Tyr Thr Val Pro Pro Lys Asn Gly Lys Ile Asn 260 265 270 Val Arg Pro Ala Glu Phe Ile Thr Arg Phe Ala Met Asn Gly Lys Phe 275 280 285 Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln 290 295 300 Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His 305 310 315 320 Ser Asn Leu Ser Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys 325 330 335 Ile Leu Thr Asp Ser Tyr Lys Phe Arg Val Lys Asp Gly Ser Phe Val 340 345 350 Thr Leu Lys Ser Lys Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Lys 355 360 365 Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly Arg Ser Glu 370 375 380 Thr Ala Val Ser Val Pro Gln Cys Arg Ser Ser Gln Ser Ser Glu Asp 385 390 395 400 Ser Phe Arg Gln Pro Cys Val Ser Val Pro Gly Ile Ser Thr Gly Thr 405 410 415 Leu Leu Gly Thr Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Val Leu 420 425 430 Ser Leu Gln Arg Ser His Ser Ser Ser Pro Glu Asp Ala Asn Pro Ser 435 440 445 Gly Val Val Arg Asp Lys His Ser Val Asn Phe Gly Ser Ala Pro Val 450 455 460 Pro Val Ser Thr Gly Glu Leu Phe Ala Leu Ser Pro Glu Thr Glu Gly 465 470 475 480 Leu Glu Ala Ala Arg Gln His Gln Ser Ser Glu Pro Ala His Cys His 485 490 495 Lys Pro Leu Leu Ser Asp Ser Thr Gln Leu Gly Phe Asp Ala Leu Cys 500 505 510 Asp Ser Asp Asp Thr Ala Met Ala Thr Phe Met Asn Tyr Leu Glu Ala 515 520 525 Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe 530 535 19 1655 DNA Rattus norvegicus CDS (33)..(1655) 19 tcacagctcc tggacctgtc agctctcttg ca atg gag ttg cca agg aaa cgt 53 Met Glu Leu Pro Arg Lys Arg 1 5 aga aga agt gat tca gag ctg ctc cag tca gaa ttc agg aca gat gca 101 Arg Arg Ser Asp Ser Glu Leu Leu Gln Ser Glu Phe Arg Thr Asp Ala 10 15 20 atg gtg gaa aac ctt ccc cgg agt ccc ttt acc tct gtt ctt tca aca 149 Met Val Glu Asn Leu Pro Arg Ser Pro Phe Thr Ser Val Leu Ser Thr 25 30 35 aga aca gga gta gca gtg ccc aat ggc atc agg gaa gct cac agc cag 197 Arg Thr Gly Val Ala Val Pro Asn Gly Ile Arg Glu Ala His Ser Gln 40 45 50 55 aca gaa aag cgt cgg aga gac aag atg aac cat ctg att tgg aaa ctg 245 Thr Glu Lys Arg Arg Arg Asp Lys Met Asn His Leu Ile Trp Lys Leu 60 65 70 tca tct atg atc cct cca cac atc ccc aca gcc cac aaa ctg gac aaa 293 Ser Ser Met Ile Pro Pro His Ile Pro Thr Ala His Lys Leu Asp Lys 75 80 85 ctg agc gtc ctg agg agg gca gtg cag tac ttg agg tct cag aga ggc 341 Leu Ser Val Leu Arg Arg Ala Val Gln Tyr Leu Arg Ser Gln Arg Gly 90 95 100 atg aca gag ttt tat tta gga gaa aat gct aaa cct tca ttt att cag 389 Met Thr Glu Phe Tyr Leu Gly Glu Asn Ala Lys Pro Ser Phe Ile Gln 105 110 115 gat aag gaa ctc agc cac tta atc ctc aag gct agt ttg atg gga cag 437 Asp Lys Glu Leu Ser His Leu Ile Leu Lys Ala Ser Leu Met Gly Gln 120 125 130 135 aac ttg ttt gac ttc tta cac cca aaa gat gtc gcc aaa gta aag gaa 485 Asn Leu Phe Asp Phe Leu His Pro Lys Asp Val Ala Lys Val Lys Glu 140 145 150 caa ctt tct tgt gat gtt tca ctg aga gag aaa ccc ata ggc acc aaa 533 Gln Leu Ser Cys Asp Val Ser Leu Arg Glu Lys Pro Ile Gly Thr Lys 155 160 165 acc tct cct cag gtt cac agt cac tcc cat att ggg cga tca cgc gtg 581 Thr Ser Pro Gln Val His Ser His Ser His Ile Gly Arg Ser Arg Val 170 175 180 cat tct ggc tcc aga cga tct ttc ttc ttt aga atg aag agc agc tgt 629 His Ser Gly Ser Arg Arg Ser Phe Phe Phe Arg Met Lys Ser Ser Cys 185 190 195 aca gtc ccc gtc aaa gaa gag caa cga tgc tcg tcc tgt tca aag aag 677 Thr Val Pro Val Lys Glu Glu Gln Arg Cys Ser Ser Cys Ser Lys Lys 200 205 210 215 aaa gac cag aga aaa ttc cac acc atc cat tgc act gga tac ttg aga 725 Lys Asp Gln Arg Lys Phe His Thr Ile His Cys Thr Gly Tyr Leu Arg 220 225 230 agc tgg cca ccg aat gtt gtg ggc acg gag aaa gag atg ggc agt ggg 773 Ser Trp Pro Pro Asn Val Val Gly Thr Glu Lys Glu Met Gly Ser Gly 235 240 245 aaa gac agt ggt cct ctt acc tgc ctt gtg gct atg gga cgg tta cag 821 Lys Asp Ser Gly Pro Leu Thr Cys Leu Val Ala Met Gly Arg Leu Gln 250 255 260 cca tat act gtc ccc ccg aag aat ggc aag atc aac gtg aga ccg gct 869 Pro Tyr Thr Val Pro Pro Lys Asn Gly Lys Ile Asn Val Arg Pro Ala 265 270 275 gag ttc ata acc cga ttc gca atg aac ggg aaa ttc gtc tac gtc gac 917 Glu Phe Ile Thr Arg Phe Ala Met Asn Gly Lys Phe Val Tyr Val Asp 280 285 290 295 caa agg gca aca gca att tta gga tac ctg cct cag gaa ctt ttg gga 965 Gln Arg Ala Thr Ala Ile Leu Gly Tyr Leu Pro Gln Glu Leu Leu Gly 300 305 310 act tcg tgt tat gaa tat ttt cat cag gat gac cac agt aat ttg agt 1013 Thr Ser Cys Tyr Glu Tyr Phe His Gln Asp Asp His Ser Asn Leu Ser 315 320 325 gac aag cac aaa gca gtt ctg cag agt aag gag aaa ata ctt aca gat 1061 Asp Lys His Lys Ala Val Leu Gln Ser Lys Glu Lys Ile Leu Thr Asp 330 335 340 tca tac aaa ttc aga gtg aag gat ggc tcc ttt gtg act ctg aag agc 1109 Ser Tyr Lys Phe Arg Val Lys Asp Gly Ser Phe Val Thr Leu Lys Ser 345 350 355 aag tgg ttc agc ttc act aac cct tgg acc aaa aag ctg gag tac atc 1157 Lys Trp Phe Ser Phe Thr Asn Pro Trp Thr Lys Lys Leu Glu Tyr Ile 360 365 370 375 gtg tct gtc aac acg ctg gtt ttg ggg cgc agt gag acc gca gta tcc 1205 Val Ser Val Asn Thr Leu Val Leu Gly Arg Ser Glu Thr Ala Val Ser 380 385 390 gtg cct cag tgc cgc agc agc cag tcc tct gaa gac tca ttt aga caa 1253 Val Pro Gln Cys Arg Ser Ser Gln Ser Ser Glu Asp Ser Phe Arg Gln 395 400 405 ccc tgc gtc agt gtg ccg ggc ata tcc aca ggg acc tta ctt ggc act 1301 Pro Cys Val Ser Val Pro Gly Ile Ser Thr Gly Thr Leu Leu Gly Thr 410 415 420 ggg agt att gga aca gat att gca aat gag gtt ctg agt tta cag agg 1349 Gly Ser Ile Gly Thr Asp Ile Ala Asn Glu Val Leu Ser Leu Gln Arg 425 430 435 tca cac tct tca tcc cca gaa gac gca aac cct tca gga gta gtg aga 1397 Ser His Ser Ser Ser Pro Glu Asp Ala Asn Pro Ser Gly Val Val Arg 440 445 450 455 gat aag cac agt gta aac ttc ggg agc gcc cct gtg ccc gtg tcc act 1445 Asp Lys His Ser Val Asn Phe Gly Ser Ala Pro Val Pro Val Ser Thr 460 465 470 ggg gag ctc ttt gca ctg agt cct gaa aca gag ggc ctg gag gct gcc 1493 Gly Glu Leu Phe Ala Leu Ser Pro Glu Thr Glu Gly Leu Glu Ala Ala 475 480 485 agg caa cac cag agt tct gag ccc gcc cac tgt cac aaa cca ctc ctc 1541 Arg Gln His Gln Ser Ser Glu Pro Ala His Cys His Lys Pro Leu Leu 490 495 500 agt gac agt acc cag ttg ggt ttt gat gcc ctg tgt gac agc gac gac 1589 Ser Asp Ser Thr Gln Leu Gly Phe Asp Ala Leu Cys Asp Ser Asp Asp 505 510 515 aca gcc atg gct aca ttc atg aat tac ctc gaa gca gag ggt ggc ctg 1637 Thr Ala Met Ala Thr Phe Met Asn Tyr Leu Glu Ala Glu Gly Gly Leu 520 525 530 535 ggt gac cct ggg gac ttc 1655 Gly Asp Pro Gly Asp Phe 540 20 541 PRT Rattus norvegicus 20 Met Glu Leu Pro Arg Lys Arg Arg Arg Ser Asp Ser Glu Leu Leu Gln 1 5 10 15 Ser Glu Phe Arg Thr Asp Ala Met Val Glu Asn Leu Pro Arg Ser Pro 20 25 30 Phe Thr Ser Val Leu Ser Thr Arg Thr Gly Val Ala Val Pro Asn Gly 35 40 45 Ile Arg Glu Ala His Ser Gln Thr Glu Lys Arg Arg Arg Asp Lys Met 50 55 60 Asn His Leu Ile Trp Lys Leu Ser Ser Met Ile Pro Pro His Ile Pro 65 70 75 80 Thr Ala His Lys Leu Asp Lys Leu Ser Val Leu Arg Arg Ala Val Gln 85 90 95 Tyr Leu Arg Ser Gln Arg Gly Met Thr Glu Phe Tyr Leu Gly Glu Asn 100 105 110 Ala Lys Pro Ser Phe Ile Gln Asp Lys Glu Leu Ser His Leu Ile Leu 115 120 125 Lys Ala Ser Leu Met Gly Gln Asn Leu Phe Asp Phe Leu His Pro Lys 130 135 140 Asp Val Ala Lys Val Lys Glu Gln Leu Ser Cys Asp Val Ser Leu Arg 145 150 155 160 Glu Lys Pro Ile Gly Thr Lys Thr Ser Pro Gln Val His Ser His Ser 165 170 175 His Ile Gly Arg Ser Arg Val His Ser Gly Ser Arg Arg Ser Phe Phe 180 185 190 Phe Arg Met Lys Ser Ser Cys Thr Val Pro Val Lys Glu Glu Gln Arg 195 200 205 Cys Ser Ser Cys Ser Lys Lys Lys Asp Gln Arg Lys Phe His Thr Ile 210 215 220 His Cys Thr Gly Tyr Leu Arg Ser Trp Pro Pro Asn Val Val Gly Thr 225 230 235 240 Glu Lys Glu Met Gly Ser Gly Lys Asp Ser Gly Pro Leu Thr Cys Leu 245 250 255 Val Ala Met Gly Arg Leu Gln Pro Tyr Thr Val Pro Pro Lys Asn Gly 260 265 270 Lys Ile Asn Val Arg Pro Ala Glu Phe Ile Thr Arg Phe Ala Met Asn 275 280 285 Gly Lys Phe Val Tyr Val Asp Gln Arg Ala Thr Ala Ile Leu Gly Tyr 290 295 300 Leu Pro Gln Glu Leu Leu Gly Thr Ser Cys Tyr Glu Tyr Phe His Gln 305 310 315 320 Asp Asp His Ser Asn Leu Ser Asp Lys His Lys Ala Val Leu Gln Ser 325 330 335 Lys Glu Lys Ile Leu Thr Asp Ser Tyr Lys Phe Arg Val Lys Asp Gly 340 345 350 Ser Phe Val Thr Leu Lys Ser Lys Trp Phe Ser Phe Thr Asn Pro Trp 355 360 365 Thr Lys Lys Leu Glu Tyr Ile Val Ser Val Asn Thr Leu Val Leu Gly 370 375 380 Arg Ser Glu Thr Ala Val Ser Val Pro Gln Cys Arg Ser Ser Gln Ser 385 390 395 400 Ser Glu Asp Ser Phe Arg Gln Pro Cys Val Ser Val Pro Gly Ile Ser 405 410 415 Thr Gly Thr Leu Leu Gly Thr Gly Ser Ile Gly Thr Asp Ile Ala Asn 420 425 430 Glu Val Leu Ser Leu Gln Arg Ser His Ser Ser Ser Pro Glu Asp Ala 435 440 445 Asn Pro Ser Gly Val Val Arg Asp Lys His Ser Val Asn Phe Gly Ser 450 455 460 Ala Pro Val Pro Val Ser Thr Gly Glu Leu Phe Ala Leu Ser Pro Glu 465 470 475 480 Thr Glu Gly Leu Glu Ala Ala Arg Gln His Gln Ser Ser Glu Pro Ala 485 490 495 His Cys His Lys Pro Leu Leu Ser Asp Ser Thr Gln Leu Gly Phe Asp 500 505 510 Ala Leu Cys Asp Ser Asp Asp Thr Ala Met Ala Thr Phe Met Asn Tyr 515 520 525 Leu Glu Ala Glu Gly Gly Leu Gly Asp Pro Gly Asp Phe 530 535 540 21 33 DNA Artificial Sequence Description of Artificial SequenceSense primer 1 21 actagtcgac ttatgttttt taccataagc acc 33 22 30 DNA Artificial Sequence Description of Artificial SequenceAntisense primer 1 22 gtcgacctgc gctactgtgg ctgagctttg 30 23 33 DNA Artificial Sequence Description of Artificial Sequenceper-F 23 cagcagatsa rctgyntsng acagyrtcmt cag 33 24 31 DNA Artificial Sequence Description of Artificial Sequenceper-R 24 gctrcactgr ctgrtgmsng acrccacrct c 31 25 24 DNA Artificial Sequence Description of Artificial SequencecPer2-R1 primer 25 ttgctgtacc aggcacatta caac 24 26 32 DNA Artificial Sequence Description of Artificial SequenceYK-F1 26 rtncaytcng gntaycargc nccnmgnatn cc 32 27 4035 DNA Gallus gallus CDS (1)..(4035) 27 atg gac tgt atc gag gtc agg ggg ttc tac tct agc act gag gag cag 48 Met Asp Cys Ile Glu Val Arg Gly Phe Tyr Ser Ser Thr Glu Glu Gln 1 5 10 15 aac cct gag cag caa gct gat atc agt gaa aac att tct tca ttg ttc 96 Asn Pro Glu Gln Gln Ala Asp Ile Ser Glu Asn Ile Ser Ser Leu Phe 20 25 30 tct tta aaa gag caa cag aaa atg agt gag tat tct gga ctt gca agt 144 Ser Leu Lys Glu Gln Gln Lys Met Ser Glu Tyr Ser Gly Leu Ala Ser 35 40 45 aac cat agc cag atg att gct gaa gat tct gaa att cag cca aaa cct 192 Asn His Ser Gln Met Ile Ala Glu Asp Ser Glu Ile Gln Pro Lys Pro 50 55 60 gaa cac tct ccc gaa gtc ctt cag gaa gat att gag atg agc agc gga 240 Glu His Ser Pro Glu Val Leu Gln Glu Asp Ile Glu Met Ser Ser Gly 65 70 75 80 tcc agt gga aat gac ttc agt gga aat gag acg aat gaa aac tac tcc 288 Ser Ser Gly Asn Asp Phe Ser Gly Asn Glu Thr Asn Glu Asn Tyr Ser 85 90 95 agt gga cat gat tct cat ggc cac gaa tct gat gaa aat ggg aaa gat 336 Ser Gly His Asp Ser His Gly His Glu Ser Asp Glu Asn Gly Lys Asp 100 105 110 tca gca atg ctc atg gaa tct tca gac tgt cat aaa agt tca agc tca 384 Ser Ala Met Leu Met Glu Ser Ser Asp Cys His Lys Ser Ser Ser Ser 115 120 125 aat gca ttt agt ctg atg att gcg aac tct gaa cac aat cag tct agc 432 Asn Ala Phe Ser Leu Met Ile Ala Asn Ser Glu His Asn Gln Ser Ser 130 135 140 agt gga tgc agc agc gag cag tct act aaa gcc aaa acg caa aag gaa 480 Ser Gly Cys Ser Ser Glu Gln Ser Thr Lys Ala Lys Thr Gln Lys Glu 145 150 155 160 ttg ttg aag aca ttg caa gag ctg aaa gct cac ctt cct gct gaa aaa 528 Leu Leu Lys Thr Leu Gln Glu Leu Lys Ala His Leu Pro Ala Glu Lys 165 170 175 aga att aaa ggc aaa tcc agt gtc cta aca aca ctg aaa tat gcc ctt 576 Arg Ile Lys Gly Lys Ser Ser Val Leu Thr Thr Leu Lys Tyr Ala Leu 180 185 190 aaa agc att aaa caa gtt aaa gcc aat gag gaa tat tac caa ttg ttg 624 Lys Ser Ile Lys Gln Val Lys Ala Asn Glu Glu Tyr Tyr Gln Leu Leu 195 200 205 atg att aat gaa tcc cag cct tct gga ctc aat gtg tca tct tat aca 672 Met Ile Asn Glu Ser Gln Pro Ser Gly Leu Asn Val Ser Ser Tyr Thr 210 215 220 gtg gaa gaa gtt gag act ata acc tca gaa tac atc atg aaa aat gca 720 Val Glu Glu Val Glu Thr Ile Thr Ser Glu Tyr Ile Met Lys Asn Ala 225 230 235 240 gat atg ttt gct gta gct gtt tct ttg att act ggg aaa att gtg tac 768 Asp Met Phe Ala Val Ala Val Ser Leu Ile Thr Gly Lys Ile Val Tyr 245 250 255 atc tct gat caa gct gct gct att ctg cgc tgt aag agg agt tat ttt 816 Ile Ser Asp Gln Ala Ala Ala Ile Leu Arg Cys Lys Arg Ser Tyr Phe 260 265 270 aaa aat gcc aaa ttt gtg gag tta ttg gca cct caa gat gtc agt gtt 864 Lys Asn Ala Lys Phe Val Glu Leu Leu Ala Pro Gln Asp Val Ser Val 275 280 285 ttc tat act tct act acc cca tac aga tta cca tct tgg aat att tgc 912 Phe Tyr Thr Ser Thr Thr Pro Tyr Arg Leu Pro Ser Trp Asn Ile Cys 290 295 300 agc aga gct gag tct tcc acc cag gat tgc atg gaa gag aaa tcc ttt 960 Ser Arg Ala Glu Ser Ser Thr Gln Asp Cys Met Glu Glu Lys Ser Phe 305 310 315 320 ttc tgt cgc atc agt gca gga aag gag cgt gaa aat gag att tgc tat 1008 Phe Cys Arg Ile Ser Ala Gly Lys Glu Arg Glu Asn Glu Ile Cys Tyr 325 330 335 cac cca ttt cgg atg act cct tac ctt atc aaa gta caa gat cca gaa 1056 His Pro Phe Arg Met Thr Pro Tyr Leu Ile Lys Val Gln Asp Pro Glu 340 345 350 gta gca gag gac caa ctt tgt tgt gtg ctc ctt gca gaa aaa gtg cac 1104 Val Ala Glu Asp Gln Leu Cys Cys Val Leu Leu Ala Glu Lys Val His 355 360 365 tct ggt tat gaa gca ccc aga att cct cca gac aaa aga att ttt aca 1152 Ser Gly Tyr Glu Ala Pro Arg Ile Pro Pro Asp Lys Arg Ile Phe Thr 370 375 380 aca aca cac aca cca acc tgt ttg ttc cag gat gta gat gag aga gct 1200 Thr Thr His Thr Pro Thr Cys Leu Phe Gln Asp Val Asp Glu Arg Ala 385 390 395 400 gta cct ctg ttg gga tac cta cct cag gac tta ata gga acg cct gtt 1248 Val Pro Leu Leu Gly Tyr Leu Pro Gln Asp Leu Ile Gly Thr Pro Val 405 410 415 ttg gtg cat ctt cac cca aat gac aga ccc tta atg cta gca att cac 1296 Leu Val His Leu His Pro Asn Asp Arg Pro Leu Met Leu Ala Ile His 420 425 430 aaa aaa ata ctt caa tat gga gga cag cct ttt gac tat tca cca atc 1344 Lys Lys Ile Leu Gln Tyr Gly Gly Gln Pro Phe Asp Tyr Ser Pro Ile 435 440 445 agg ttt tgc act aga aat gga gat tat ata acc atg gac act agc tgg 1392 Arg Phe Cys Thr Arg Asn Gly Asp Tyr Ile Thr Met Asp Thr Ser Trp 450 455 460 tcc agt ttc atc aat cct tgg agt cga aag gtt tca ttt atc att gga 1440 Ser Ser Phe Ile Asn Pro Trp Ser Arg Lys Val Ser Phe Ile Ile Gly 465 470 475 480 aga cac aaa gtt agg acg ggt ccc tta aat gaa gat gtt ttt gcc gct 1488 Arg His Lys Val Arg Thr Gly Pro Leu Asn Glu Asp Val Phe Ala Ala 485 490 495 ccc aac tat acg gag gac aga atc ctt cac ccc agt gtt cag gag atc 1536 Pro Asn Tyr Thr Glu Asp Arg Ile Leu His Pro Ser Val Gln Glu Ile 500 505 510 aca gag caa ata tat cgg ctg tta cta cag cct gta cac aac agt gga 1584 Thr Glu Gln Ile Tyr Arg Leu Leu Leu Gln Pro Val His Asn Ser Gly 515 520 525 tcc agt ggc tat gga agt cta ggt agc aat ggc tca cac gaa cac tta 1632 Ser Ser Gly Tyr Gly Ser Leu Gly Ser Asn Gly Ser His Glu His Leu 530 535 540 atg agt gtg gca tcc tcc agt gac agc aca gga aat aat aat gat gac 1680 Met Ser Val Ala Ser Ser Ser Asp Ser Thr Gly Asn Asn Asn Asp Asp 545 550 555 560 aca caa aag gat aaa aca ata agt caa gat gcc cgt aag gtc aaa act 1728 Thr Gln Lys Asp Lys Thr Ile Ser Gln Asp Ala Arg Lys Val Lys Thr 565 570 575 aaa gga cag cat att ttc act gag aat aaa gga aaa ctg gaa tat aaa 1776 Lys Gly Gln His Ile Phe Thr Glu Asn Lys Gly Lys Leu Glu Tyr Lys 580 585 590 aga gag cct tct gca gaa aaa caa aat ggt cct ggt ggt cag gtg aaa 1824 Arg Glu Pro Ser Ala Glu Lys Gln Asn Gly Pro Gly Gly Gln Val Lys 595 600 605 gat gtg ata gga aag gat acc aca gct aca gct gct cct aaa aat gtg 1872 Asp Val Ile Gly Lys Asp Thr Thr Ala Thr Ala Ala Pro Lys Asn Val 610 615 620 gct act gaa gag ttg gca tgg aaa gaa caa cct gta tat tct tat caa 1920 Ala Thr Glu Glu Leu Ala Trp Lys Glu Gln Pro Val Tyr Ser Tyr Gln 625 630 635 640 cag att agc tgc ttg gat agt gtc atc agg tat ttg gag agt tgt aat 1968 Gln Ile Ser Cys Leu Asp Ser Val Ile Arg Tyr Leu Glu Ser Cys Asn 645 650 655 gtg cct ggt aca gca aaa aga aaa tgt gaa cct tca tca agt gtg aat 2016 Val Pro Gly Thr Ala Lys Arg Lys Cys Glu Pro Ser Ser Ser Val Asn 660 665 670 tct agt gtt cac gag caa aaa gca tct gtt aat gct ata caa ccc tta 2064 Ser Ser Val His Glu Gln Lys Ala Ser Val Asn Ala Ile Gln Pro Leu 675 680 685 gga gac tct act gtg ttg aag tca tct ggt aaa tca agt ggt ccc cca 2112 Gly Asp Ser Thr Val Leu Lys Ser Ser Gly Lys Ser Ser Gly Pro Pro 690 695 700 gta gtt ggt gct cac tta act tct ttg gcc tta cct ggc aag cct gaa 2160 Val Val Gly Ala His Leu Thr Ser Leu Ala Leu Pro Gly Lys Pro Glu 705 710 715 720 agt gtt gta tcg ctc acc agt cag tgc agc tac agt agc acc att gtt 2208 Ser Val Val Ser Leu Thr Ser Gln Cys Ser Tyr Ser Ser Thr Ile Val 725 730 735 cat gtt gga gac aaa aaa cca caa cct gaa tta gaa atg ata gaa gat 2256 His Val Gly Asp Lys Lys Pro Gln Pro Glu Leu Glu Met Ile Glu Asp 740 745 750 ggt cca agt gga gca gaa gtc tta gat act caa ctt cct gcc cct cca 2304 Gly Pro Ser Gly Ala Glu Val Leu Asp Thr Gln Leu Pro Ala Pro Pro 755 760 765 ccc agc tct acg cat gta aat cag gaa aag gag tca ttt aaa aaa ctg 2352 Pro Ser Ser Thr His Val Asn Gln Glu Lys Glu Ser Phe Lys Lys Leu 770 775 780 gga ctt aca aag gaa gtc ctt gca gtg cat aca caa aaa gaa gag caa 2400 Gly Leu Thr Lys Glu Val Leu Ala Val His Thr Gln Lys Glu Glu Gln 785 790 795 800 agc ttt ttg aat aag ttc aaa gaa atc aag aga ttc aat att ttc cag 2448 Ser Phe Leu Asn Lys Phe Lys Glu Ile Lys Arg Phe Asn Ile Phe Gln 805 810 815 tcc cac tgc aat tac tac tta caa gat aaa cct aaa gga agg cct ggt 2496 Ser His Cys Asn Tyr Tyr Leu Gln Asp Lys Pro Lys Gly Arg Pro Gly 820 825 830 gaa cgt ggt ggc cgc gga caa cga aat gga act tct gga atg gat cag 2544 Glu Arg Gly Gly Arg Gly Gln Arg Asn Gly Thr Ser Gly Met Asp Gln 835 840 845 cct tgg aag aaa agt ggg aaa aac agg aaa tca aaa cgc att aaa cca 2592 Pro Trp Lys Lys Ser Gly Lys Asn Arg Lys Ser Lys Arg Ile Lys Pro 850 855 860 cag gag tct tca gac agt aca act tct gga act aaa ttc ccc cat cgc 2640 Gln Glu Ser Ser Asp Ser Thr Thr Ser Gly Thr Lys Phe Pro His Arg 865 870 875 880 ttt cct ctt cag ggt tta aat act acc gct tgg tca ccg tca gac act 2688 Phe Pro Leu Gln Gly Leu Asn Thr Thr Ala Trp Ser Pro Ser Asp Thr 885 890 895 tca caa gca agc tac tca gcg atg tct ttt cca act gtt atg cct gca 2736 Ser Gln Ala Ser Tyr Ser Ala Met Ser Phe Pro Thr Val Met Pro Ala 900 905 910 tat ccg ctt cct gtt ttt cca gca gca gca gga act gtg cca cca gct 2784 Tyr Pro Leu Pro Val Phe Pro Ala Ala Ala Gly Thr Val Pro Pro Ala 915 920 925 cct gag act tca gtc tct ggt ttt aat cag ctg cca gac tcg gga aat 2832 Pro Glu Thr Ser Val Ser Gly Phe Asn Gln Leu Pro Asp Ser Gly Asn 930 935 940 act tgc tct atg caa cca tcc cag ttt tct gcc cct ctt atg aca ccc 2880 Thr Cys Ser Met Gln Pro Ser Gln Phe Ser Ala Pro Leu Met Thr Pro 945 950 955 960 gtt gta gct ctt gtg ctc ccc aac tat gtc tac cca gaa atg aac aat 2928 Val Val Ala Leu Val Leu Pro Asn Tyr Val Tyr Pro Glu Met Asn Asn 965 970 975 agc tta cct caa aca ctt tac cac agc caa gcc aat ttt ccc acc cat 2976 Ser Leu Pro Gln Thr Leu Tyr His Ser Gln Ala Asn Phe Pro Thr His 980 985 990 cct gct ttc tct tca cag aca gta ttt cca gcg cag cct cca ttc act 3024 Pro Ala Phe Ser Ser Gln Thr Val Phe Pro Ala Gln Pro Pro Phe Thr 995 1000 1005 acc cct agc cct ttc cca caa cag gcg ttt ttt cca atg caa cca ttc 3072 Thr Pro Ser Pro Phe Pro Gln Gln Ala Phe Phe Pro Met Gln Pro Phe 1010 1015 1020 cat tat aat cca cca gca gaa att gaa aag gtt cct gtc aca gag aca 3120 His Tyr Asn Pro Pro Ala Glu Ile Glu Lys Val Pro Val Thr Glu Thr 1025 1030 1035 1040 cga aac gag cca tcc cgt tcc tgc act cca cag tca gtg ggt cct caa 3168 Arg Asn Glu Pro Ser Arg Ser Cys Thr Pro Gln Ser Val Gly Pro Gln 1045 1050 1055 gac cag gct tca ccg cct ttg ttc caa tca agg tgt agt tct cct ctg 3216 Asp Gln Ala Ser Pro Pro Leu Phe Gln Ser Arg Cys Ser Ser Pro Leu 1060 1065 1070 aat ctt cta cag ttg gaa gaa aac aca aaa act gtg gaa agt gga gct 3264 Asn Leu Leu Gln Leu Glu Glu Asn Thr Lys Thr Val Glu Ser Gly Ala 1075 1080 1085 cct gca ggt ttg cat gga gct tta aat gag gaa gga acc ata ggc aaa 3312 Pro Ala Gly Leu His Gly Ala Leu Asn Glu Glu Gly Thr Ile Gly Lys 1090 1095 1100 atc atg aca act gat gct ggt agt gga aag gga tcc cta cca gct gag 3360 Ile Met Thr Thr Asp Ala Gly Ser Gly Lys Gly Ser Leu Pro Ala Glu 1105 1110 1115 1120 tct cca atg gat gct caa aat agc gat gca ctc tcc atg tcc agt gtc 3408 Ser Pro Met Asp Ala Gln Asn Ser Asp Ala Leu Ser Met Ser Ser Val 1125 1130 1135 ctg ctt gac att tta ctt caa gaa gat gca tgc tca ggc act ggc tca 3456 Leu Leu Asp Ile Leu Leu Gln Glu Asp Ala Cys Ser Gly Thr Gly Ser 1140 1145 1150 gct tcc tca ggg agc ggt gta tct gca gct gct gaa tct ctc ggg tct 3504 Ala Ser Ser Gly Ser Gly Val Ser Ala Ala Ala Glu Ser Leu Gly Ser 1155 1160 1165 gga tct aac ggc tgt gac atg tca ggg agc aga aca ggc agt agt gaa 3552 Gly Ser Asn Gly Cys Asp Met Ser Gly Ser Arg Thr Gly Ser Ser Glu 1170 1175 1180 act agt cat acc agc aaa tac ttt ggg agt atc gat tct tca gaa aac 3600 Thr Ser His Thr Ser Lys Tyr Phe Gly Ser Ile Asp Ser Ser Glu Asn 1185 1190 1195 1200 cat cat aaa aca aaa atg aag gca gaa ata gaa gaa agt gag cac ttc 3648 His His Lys Thr Lys Met Lys Ala Glu Ile Glu Glu Ser Glu His Phe 1205 1210 1215 att aaa tat gtt ctt cag gat cct ata tgg ctt ttg atg gca aac aca 3696 Ile Lys Tyr Val Leu Gln Asp Pro Ile Trp Leu Leu Met Ala Asn Thr 1220 1225 1230 gat gac acc gtt atg atg act tac cag tta ccc tct aga gat ttg gaa 3744 Asp Asp Thr Val Met Met Thr Tyr Gln Leu Pro Ser Arg Asp Leu Glu 1235 1240 1245 aca gtt tta aaa gaa gat aag ctg aaa cta aag caa atg cag aaa cta 3792 Thr Val Leu Lys Glu Asp Lys Leu Lys Leu Lys Gln Met Gln Lys Leu 1250 1255 1260 caa cca aaa ttt act gaa gac caa aaa aga gag ctt att gaa gtt cat 3840 Gln Pro Lys Phe Thr Glu Asp Gln Lys Arg Glu Leu Ile Glu Val His 1265 1270 1275 1280 cca tgg atc cag caa ggt gga ctg cca aag act gtt gct aac tct gaa 3888 Pro Trp Ile Gln Gln Gly Gly Leu Pro Lys Thr Val Ala Asn Ser Glu 1285 1290 1295 tgt att ttt tgt gag gac aat ata cag agc aat ttt tat aca tcg tac 3936 Cys Ile Phe Cys Glu Asp Asn Ile Gln Ser Asn Phe Tyr Thr Ser Tyr 1300 1305 1310 gat gaa gaa atc cat gaa atg gac ctt aat gaa atg att gaa gac agt 3984 Asp Glu Glu Ile His Glu Met Asp Leu Asn Glu Met Ile Glu Asp Ser 1315 1320 1325 ggg gag aac aat ttg gtt cct ctg agt caa gtc aat gaa gaa caa aca 4032 Gly Glu Asn Asn Leu Val Pro Leu Ser Gln Val Asn Glu Glu Gln Thr 1330 1335 1340 tag 4035 28 1344 PRT Gallus gallus 28 Met Asp Cys Ile Glu Val Arg Gly Phe Tyr Ser Ser Thr Glu Glu Gln 1 5 10 15 Asn Pro Glu Gln Gln Ala Asp Ile Ser Glu Asn Ile Ser Ser Leu Phe 20 25 30 Ser Leu Lys Glu Gln Gln Lys Met Ser Glu Tyr Ser Gly Leu Ala Ser 35 40 45 Asn His Ser Gln Met Ile Ala Glu Asp Ser Glu Ile Gln Pro Lys Pro 50 55 60 Glu His Ser Pro Glu Val Leu Gln Glu Asp Ile Glu Met Ser Ser Gly 65 70 75 80 Ser Ser Gly Asn Asp Phe Ser Gly Asn Glu Thr Asn Glu Asn Tyr Ser 85 90 95 Ser Gly His Asp Ser His Gly His Glu Ser Asp Glu Asn Gly Lys Asp 100 105 110 Ser Ala Met Leu Met Glu Ser Ser Asp Cys His Lys Ser Ser Ser Ser 115 120 125 Asn Ala Phe Ser Leu Met Ile Ala Asn Ser Glu His Asn Gln Ser Ser 130 135 140 Ser Gly Cys Ser Ser Glu Gln Ser Thr Lys Ala Lys Thr Gln Lys Glu 145 150 155 160 Leu Leu Lys Thr Leu Gln Glu Leu Lys Ala His Leu Pro Ala Glu Lys 165 170 175 Arg Ile Lys Gly Lys Ser Ser Val Leu Thr Thr Leu Lys Tyr Ala Leu 180 185 190 Lys Ser Ile Lys Gln Val Lys Ala Asn Glu Glu Tyr Tyr Gln Leu Leu 195 200 205 Met Ile Asn Glu Ser Gln Pro Ser Gly Leu Asn Val Ser Ser Tyr Thr 210 215 220 Val Glu Glu Val Glu Thr Ile Thr Ser Glu Tyr Ile Met Lys Asn Ala 225 230 235 240 Asp Met Phe Ala Val Ala Val Ser Leu Ile Thr Gly Lys Ile Val Tyr 245 250 255 Ile Ser Asp Gln Ala Ala Ala Ile Leu Arg Cys Lys Arg Ser Tyr Phe 260 265 270 Lys Asn Ala Lys Phe Val Glu Leu Leu Ala Pro Gln Asp Val Ser Val 275 280 285 Phe Tyr Thr Ser Thr Thr Pro Tyr Arg Leu Pro Ser Trp Asn Ile Cys 290 295 300 Ser Arg Ala Glu Ser Ser Thr Gln Asp Cys Met Glu Glu Lys Ser Phe 305 310 315 320 Phe Cys Arg Ile Ser Ala Gly Lys Glu Arg Glu Asn Glu Ile Cys Tyr 325 330 335 His Pro Phe Arg Met Thr Pro Tyr Leu Ile Lys Val Gln Asp Pro Glu 340 345 350 Val Ala Glu Asp Gln Leu Cys Cys Val Leu Leu Ala Glu Lys Val His 355 360 365 Ser Gly Tyr Glu Ala Pro Arg Ile Pro Pro Asp Lys Arg Ile Phe Thr 370 375 380 Thr Thr His Thr Pro Thr Cys Leu Phe Gln Asp Val Asp Glu Arg Ala 385 390 395 400 Val Pro Leu Leu Gly Tyr Leu Pro Gln Asp Leu Ile Gly Thr Pro Val 405 410 415 Leu Val His Leu His Pro Asn Asp Arg Pro Leu Met Leu Ala Ile His 420 425 430 Lys Lys Ile Leu Gln Tyr Gly Gly Gln Pro Phe Asp Tyr Ser Pro Ile 435 440 445 Arg Phe Cys Thr Arg Asn Gly Asp Tyr Ile Thr Met Asp Thr Ser Trp 450 455 460 Ser Ser Phe Ile Asn Pro Trp Ser Arg Lys Val Ser Phe Ile Ile Gly 465 470 475 480 Arg His Lys Val Arg Thr Gly Pro Leu Asn Glu Asp Val Phe Ala Ala 485 490 495 Pro Asn Tyr Thr Glu Asp Arg Ile Leu His Pro Ser Val Gln Glu Ile 500 505 510 Thr Glu Gln Ile Tyr Arg Leu Leu Leu Gln Pro Val His Asn Ser Gly 515 520 525 Ser Ser Gly Tyr Gly Ser Leu Gly Ser Asn Gly Ser His Glu His Leu 530 535 540 Met Ser Val Ala Ser Ser Ser Asp Ser Thr Gly Asn Asn Asn Asp Asp 545 550 555 560 Thr Gln Lys Asp Lys Thr Ile Ser Gln Asp Ala Arg Lys Val Lys Thr 565 570 575 Lys Gly Gln His Ile Phe Thr Glu Asn Lys Gly Lys Leu Glu Tyr Lys 580 585 590 Arg Glu Pro Ser Ala Glu Lys Gln Asn Gly Pro Gly Gly Gln Val Lys 595 600 605 Asp Val Ile Gly Lys Asp Thr Thr Ala Thr Ala Ala Pro Lys Asn Val 610 615 620 Ala Thr Glu Glu Leu Ala Trp Lys Glu Gln Pro Val Tyr Ser Tyr Gln 625 630 635 640 Gln Ile Ser Cys Leu Asp Ser Val Ile Arg Tyr Leu Glu Ser Cys Asn 645 650 655 Val Pro Gly Thr Ala Lys Arg Lys Cys Glu Pro Ser Ser Ser Val Asn 660 665 670 Ser Ser Val His Glu Gln Lys Ala Ser Val Asn Ala Ile Gln Pro Leu 675 680 685 Gly Asp Ser Thr Val Leu Lys Ser Ser Gly Lys Ser Ser Gly Pro Pro 690 695 700 Val Val Gly Ala His Leu Thr Ser Leu Ala Leu Pro Gly Lys Pro Glu 705 710 715 720 Ser Val Val Ser Leu Thr Ser Gln Cys Ser Tyr Ser Ser Thr Ile Val 725 730 735 His Val Gly Asp Lys Lys Pro Gln Pro Glu Leu Glu Met Ile Glu Asp 740 745 750 Gly Pro Ser Gly Ala Glu Val Leu Asp Thr Gln Leu Pro Ala Pro Pro 755 760 765 Pro Ser Ser Thr His Val Asn Gln Glu Lys Glu Ser Phe Lys Lys Leu 770 775 780 Gly Leu Thr Lys Glu Val Leu Ala Val His Thr Gln Lys Glu Glu Gln 785 790 795 800 Ser Phe Leu Asn Lys Phe Lys Glu Ile Lys Arg Phe Asn Ile Phe Gln 805 810 815 Ser His Cys Asn Tyr Tyr Leu Gln Asp Lys Pro Lys Gly Arg Pro Gly 820 825 830 Glu Arg Gly Gly Arg Gly Gln Arg Asn Gly Thr Ser Gly Met Asp Gln 835 840 845 Pro Trp Lys Lys Ser Gly Lys Asn Arg Lys Ser Lys Arg Ile Lys Pro 850 855 860 Gln Glu Ser Ser Asp Ser Thr Thr Ser Gly Thr Lys Phe Pro His Arg 865 870 875 880 Phe Pro Leu Gln Gly Leu Asn Thr Thr Ala Trp Ser Pro Ser Asp Thr 885 890 895 Ser Gln Ala Ser Tyr Ser Ala Met Ser Phe Pro Thr Val Met Pro Ala 900 905 910 Tyr Pro Leu Pro Val Phe Pro Ala Ala Ala Gly Thr Val Pro Pro Ala 915 920 925 Pro Glu Thr Ser Val Ser Gly Phe Asn Gln Leu Pro Asp Ser Gly Asn 930 935 940 Thr Cys Ser Met Gln Pro Ser Gln Phe Ser Ala Pro Leu Met Thr Pro 945 950 955 960 Val Val Ala Leu Val Leu Pro Asn Tyr Val Tyr Pro Glu Met Asn Asn 965 970 975 Ser Leu Pro Gln Thr Leu Tyr His Ser Gln Ala Asn Phe Pro Thr His 980 985 990 Pro Ala Phe Ser Ser Gln Thr Val Phe Pro Ala Gln Pro Pro Phe Thr 995 1000 1005 Thr Pro Ser Pro Phe Pro Gln Gln Ala Phe Phe Pro Met Gln Pro Phe 1010 1015 1020 His Tyr Asn Pro Pro Ala Glu Ile Glu Lys Val Pro Val Thr Glu Thr 1025 1030 1035 1040 Arg Asn Glu Pro Ser Arg Ser Cys Thr Pro Gln Ser Val Gly Pro Gln 1045 1050 1055 Asp Gln Ala Ser Pro Pro Leu Phe Gln Ser Arg Cys Ser Ser Pro Leu 1060 1065 1070 Asn Leu Leu Gln Leu Glu Glu Asn Thr Lys Thr Val Glu Ser Gly Ala 1075 1080 1085 Pro Ala Gly Leu His Gly Ala Leu Asn Glu Glu Gly Thr Ile Gly Lys 1090 1095 1100 Ile Met Thr Thr Asp Ala Gly Ser Gly Lys Gly Ser Leu Pro Ala Glu 1105 1110 1115 1120 Ser Pro Met Asp Ala Gln Asn Ser Asp Ala Leu Ser Met Ser Ser Val 1125 1130 1135 Leu Leu Asp Ile Leu Leu Gln Glu Asp Ala Cys Ser Gly Thr Gly Ser 1140 1145 1150 Ala Ser Ser Gly Ser Gly Val Ser Ala Ala Ala Glu Ser Leu Gly Ser 1155 1160 1165 Gly Ser Asn Gly Cys Asp Met Ser Gly Ser Arg Thr Gly Ser Ser Glu 1170 1175 1180 Thr Ser His Thr Ser Lys Tyr Phe Gly Ser Ile Asp Ser Ser Glu Asn 1185 1190 1195 1200 His His Lys Thr Lys Met Lys Ala Glu Ile Glu Glu Ser Glu His Phe 1205 1210 1215 Ile Lys Tyr Val Leu Gln Asp Pro Ile Trp Leu Leu Met Ala Asn Thr 1220 1225 1230 Asp Asp Thr Val Met Met Thr Tyr Gln Leu Pro Ser Arg Asp Leu Glu 1235 1240 1245 Thr Val Leu Lys Glu Asp Lys Leu Lys Leu Lys Gln Met Gln Lys Leu 1250 1255 1260 Gln Pro Lys Phe Thr Glu Asp Gln Lys Arg Glu Leu Ile Glu Val His 1265 1270 1275 1280 Pro Trp Ile Gln Gln Gly Gly Leu Pro Lys Thr Val Ala Asn Ser Glu 1285 1290 1295 Cys Ile Phe Cys Glu Asp Asn Ile Gln Ser Asn Phe Tyr Thr Ser Tyr 1300 1305 1310 Asp Glu Glu Ile His Glu Met Asp Leu Asn Glu Met Ile Glu Asp Ser 1315 1320 1325 Gly Glu Asn Asn Leu Val Pro Leu Ser Gln Val Asn Glu Glu Gln Thr 1330 1335 1340 29 22 DNA Artificial Sequence Description of Artificial SequenceBMAL-F 29 gtgctmmgga tggcwgtkca gc 22 30 22 DNA Artificial Sequence Description of Artificial SequenceBMAL-R 30 gcgyccratt gcvacraggc ag 22 31 20 DNA Artificial Sequence Description of Artificial SequencehB2F1 31 gaccaagtgg ctcctgcgat 20 32 20 DNA Artificial Sequence Description of Artificial SequencehB2R1 32 gctagagggt ccactggatg 20 33 27 DNA Artificial Sequence Description of Artificial SequencehBMAL2-F4 33 gtgctggtag tattggaaca gatattg 27 34 20 DNA Artificial Sequence Description of Artificial SequencehBMAL2-R1 34 gctagagggt ccactggatg 20 35 32 DNA Artificial Sequence Description of Artificial SequencemBMAL2-F1 35 ggtcgaccac catggagttt tccaaggaaa cg 32 36 24 DNA Artificial Sequence Description of Artificial SequencemBMAL2-R1 36 gctagagtgc ccactggatg tcac 24 37 19 DNA Artificial Sequence Description of Artificial Sequence Sense primer 2 37 catgtctggc agaggcaag 19 38 19 DNA Artificial Sequence Description of Artificial SequenceAntisense primer 2 38 ttagccgccg aagccgtag 19 39 21 DNA Artificial Sequence Description of Artificial Sequence cB1F1600-primer 39 tccagacatt tcttcagctg g 21 40 19 DNA Artificial Sequence Description of Artificial Sequence cB1REND-primer 40 ggatgttgaa gcaaggtgc 19 41 21 DNA Artificial Sequence Description of Artificial Sequence cB2F1270-primer 41 acgagtactg ccatcaagat g 21 42 20 DNA Artificial Sequence Description of Artificial Sequence cB2REND-primer 42 gagagcccat tggatgtcac 20 43 19 DNA Artificial Sequence Description of Artificial Sequence cqCF862-primer 43 ttcttggatc acagggcac 19 44 22 DNA Artificial Sequence Description of Artificial Sequence cqCR1364-primer 44 ggagtgctag tgtccactgt ca 22 45 21 DNA Artificial Sequence Description of Artificial Sequence cP2RTF-primer 45 ggaagtcctt gcagtgcata c 21 46 19 DNA Artificial Sequence Description of Artificial Sequence cP2RTR-primer 46 acaggaagcg gatatgcag 19 47 20 DNA Artificial Sequence Description of Artificial SequencecGAF-primer 47 accactgtcc atgccatcac 20 48 20 DNA Artificial Sequence Description of Artificial SequencecGAR-primer 48 tccacaacac ggttgctgta 20 49 19 DNA Artificial Sequence Description of Artificial SequencemBMAL2-F2 primer 49 tggttggatg cgaaagagg 19 50 21 DNA Artificial Sequence Description of Artificial SequencemBMAL2-R4 primer 50 aggtttctct cttggtgaac c 21 51 19 DNA Artificial Sequence Description of Artificial SequencermBmal1-F1 primer 51 tggtaccaac atgcaatgc 19 52 21 DNA Artificial Sequence Description of Artificial SequencermBmal1-R1 primer 52 agtgtccgag gaagatagct g 21 53 22 DNA Artificial Sequence Description of Artificial SequencermPer2-F1 primer 53 gctcactgcc agaactatct cc 22 54 22 DNA Artificial Sequence Description of Artificial SequencermPer2-R1 primer 54 cctctagctg aagcaggtta ag 22 55 21 DNA Artificial Sequence Description of Artificial SequencermClock-F1 primer 55 caaggtcagc aacttgtgac c 21 56 20 DNA Artificial Sequence Description of Artificial SequencermClock-R1 primer 56 aggatgagct gtgtcgaagg 20 57 20 DNA Artificial Sequence Description of Artificial SequencemGAPDH F1 primer 57 catcaccatc ttccaggagc 20 58 19 DNA Artificial Sequence Description of Artificial SequencemGAPDH-R1 primer 58 attgagagca atgccagcc 19 59 19 DNA Artificial Sequence Description of Artificial SequencecP2E1-S 59 gtgtcacacg tgaggctta 19 60 19 DNA Artificial Sequence Description of Artificial SequencecP2E1-AS 60 taagcctcac gtgtgacac 19 61 29 DNA Artificial Sequence Description of Artificial SequenceSense primer 3 61 tcgagctctt tggtacctgg ccagcaacc 29 62 23 DNA Artificial Sequence Description of Artificial SequenceAntisense primer 3 62 tcacgacacc tggccgttcg agg 23 63 41 DNA Artificial Sequence Description of Artificial SequenceTRE and flanking sequence 63 cggctgactc atcaagctga ctcatcaagc tgactcatca a 41 

1. DNA encoding a protein (a) or (b) below, p1 (a) a protein comprising an amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8, (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8 and which has the BMAL2 activity.
 2. DNA containing a base sequence shown by Seq. ID No. 1, 3, 5 or 7 or its complementary sequence and part or whole of these sequences.
 3. DNA which hybridizes with DNA of claim 2 under a stringent condition and which encodes a protein having the BMAL2 activity.
 4. DNA encoding a protein (a) or (b) below, (a) a protein comprising an amino acid sequence shown by Seq. ID No. 10, (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 10 and which has the BMAL2 activity.
 5. DNA containing a base sequence shown by Seq. ID No. 9 or its complementary sequence and part or whole of these sequences.
 6. DNA which hybridizes with DNA of claim 5 under a stringent condition and which encodes a protein having the BMAL2 activity.
 7. DNA encoding a protein (a) or (b) below, (a) a protein comprising an amino acid sequence shown by Seq. ID No. 12 or 14, (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 12 or 14 and which has the BMAL2 activity.
 8. DNA containing a base sequence shown by Seq. ID No. 11 or 13 or its complementary sequence and part or whole of these sequences.
 9. DNA which hybridizes with DNA of claim 8 under a stringent condition and which encodes a protein having the BMAL2 activity.
 10. DNA encoding a protein (a) or (b) below, (a) a protein comprising an amino acid sequence shown by Seq. ID No. 16, 18 or 20, (b) a protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 16, 18 or 20 and which has the BMAL2 activity.
 11. DNA containing a base sequence shown by Seq. ID No. 15, 17 or 19 or its complementary sequence and part or whole of these sequences.
 12. DNA which hybridizes with DNA of claim 11 under a stringent condition and which encodes a protein having the BMAL2 activity.
 13. A protein comprising an amino acid sequence shown by Seq. ID No. 2, 4, 6 or
 8. 14. A protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 2, 4, 6 or 8 and which has the BMAL2 activity.
 15. A protein comprising an amino acid sequence shown by Seq. ID No.
 10. 16. A protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 10 and which has the BMAL2 activity.
 17. A protein comprising an amino acid sequence shown by Seq. ID No. 12 or
 14. 18. A protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 12 or 14 and which has the BMAL2 activity.
 19. A protein comprising an amino acid sequence shown by Seq. ID No. 16, 18 or
 20. 20. A protein which comprises an amino acid sequence wherein one or a few amino acids are deleted, substituted or added in the amino acid sequence shown by Seq. ID No. 16, 18 or 20 and which has the BMAL2 activity.
 21. A peptide which comprises part of the protein of any of claims 13-20 and which has the BMAL2 activity.
 22. A fusion protein or a fusion peptide wherein the protein of any of claims 13-20 or the peptide of claim 21 is bound with a marker protein and/or a peptide tag.
 23. An antibody which specifically binds to the protein of any of claims 13-20 or to the peptide of claim
 21. 24. The antibody according to claim 23, wherein the antibody is a monoclonal antibody.
 25. A recombinant protein or peptide to which the antibody of claim 23 or 24 specifically binds and which has the BMAL2 activity.
 26. A host cell comprising an expression system capable of expressing the protein of any of claims 13-20 or the peptide of claim
 21. 27. The host cell according to claim 26, wherein the host cell is further capable of expressing CLOCK and/or BMAL1.
 28. The host cell according to claim 26 or 27, wherein the expression system at least comprises a promoter having an E-box sequence (CACGTG).
 29. The host cell according to claim 28, wherein the promoter having an E-box sequence (CACGTG) is a promoter of Per gene, Tim gene, Cry gene, vasopressin gene or the albumin D-site binding protein gene.
 30. A non-human animal which, on its chromosome, is deficient in the gene function to encode the protein of any of claims 13-20 or the peptide of claim 21 or which over-expresses the protein of any of claims 13-20 or the peptide of claim
 21. 31. The non-human animal according to claim 30, wherein the non-human animal is a mouse or a rat.
 32. A method for screening a promoter or a suppressor for the expression of the protein of any of claims 13-20 the peptide of claim 21 or a promoter or a suppressor of the Bmal2 activity, wherein a cell expressing the protein or peptide and a test substance are used.
 33. The method for screening a promoter or a suppressor for the expression of the protein/peptide or a promoter or a suppressor of the Bmal2 activity according to claim 32, wherein the cell expressing the protein of any of claims 13-20 or the peptide of claim 21 is the host cell of any of claims 26-29.
 34. A method for screening a promoter or a suppressor for the expression of the protein of any of claims 13-20 the peptide of claim 21 or a promoter or a suppressor of the Bmal2 activity, wherein the non-human animal of claim 30 or 31 and a test substance are used.
 35. An expression promoter of the protein of any of claims 13-20 or the peptide of claim 21, wherein the expression promoter is obtained by the screening method according to any of claims 32-34.
 36. An expression suppressor for the protein of any of claims 13-20 or the peptide of claim 21, wherein the expression promoter is obtained by the screening method according to any of claims 32-34.
 37. A promoter of the Bmal2 activity obtained by the screening method according to any of claims 32-34.
 38. A suppressor for the Bmal2 activity obtained by the screening method according to any of claims 32-34.
 39. A method for screening a promoter or a suppressor for the promoter transactivation, wherein a cell which expresses the protein of any of claims 13-20 or the peptide of claim 21 and which contains a promoter having an E-box sequence (CACGTG) and a test substance are used.
 40. The method for screening a promoter or a suppressor for the promoter transactivation according to claim 39, wherein the cell which expresses the protein of any of claims 13-20 or the peptide of claim 21 and which contains a promoter having an E-box sequence (CACGTG) is the host cell of claim 28 or
 29. 41. A method for screening a promoter or a suppressor for the transactivation for a promoter having an E-box sequence (CACGTG) in the non-human animal of claim 30 or 31, wherein the non-human animal and a test substance are used.
 42. A promoter of the promoter transactivation obtained by the screening method according to any of claims 39-41.
 43. A suppressor for the promoter transactivation obtained by the screening method according to any of claims 39-41.
 44. A method for diagnosing diseases associated with the expression or the activity of BMAL2, wherein the DNA sequence encoding BMAL2 in a sample is compared with the DNA sequence encoding the protein of claim 13 or
 14. 